WO2022123968A1 - Raman analysis plate, raman analysis device, analysis system, and raman analysis method - Google Patents

Raman analysis plate, raman analysis device, analysis system, and raman analysis method Download PDF

Info

Publication number
WO2022123968A1
WO2022123968A1 PCT/JP2021/040560 JP2021040560W WO2022123968A1 WO 2022123968 A1 WO2022123968 A1 WO 2022123968A1 JP 2021040560 W JP2021040560 W JP 2021040560W WO 2022123968 A1 WO2022123968 A1 WO 2022123968A1
Authority
WO
WIPO (PCT)
Prior art keywords
raman
sample
analysis
plate
raman analysis
Prior art date
Application number
PCT/JP2021/040560
Other languages
French (fr)
Japanese (ja)
Inventor
慧 若林
皓之 森村
伸介 柏木
Original Assignee
株式会社堀場製作所
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 株式会社堀場製作所 filed Critical 株式会社堀場製作所
Priority to JP2022568109A priority Critical patent/JPWO2022123968A1/ja
Publication of WO2022123968A1 publication Critical patent/WO2022123968A1/en

Links

Images

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/80—Fraction collectors

Definitions

  • the present invention relates to a Raman analysis plate and a Raman analyzer using the plate.
  • Patent Document 1 As a Raman analyzer, as shown in Patent Document 1, there is a device configured to analyze a liquid sample separated by chromatography by Raman spectroscopy.
  • this device separates the separated liquid sample into multiple locations on a glass plate or water-repellent plate and dries it, and acquires the Raman spectrum of the dried sample.
  • the separated liquid sample moves on the plate (for example, if the sample is an organic solvent, wetting occurs and droplets spread. ), It is difficult to hold it stably.
  • the main object of the present invention is to provide a plate for Raman analysis capable of stably holding a liquid sample separated by chromatography.
  • the Raman analysis plate according to the present invention is a Raman analysis plate used for analyzing a liquid sample separated by chromatography by Raman spectroscopy, and is provided at a plurality of locations on the plate surface and described above. It is characterized by having a metal exposed region for holding a sample and a hydrophobic region provided on the surface of the plate and surrounding the metal exposed region and having a higher hydrophobicity than the metal exposed region.
  • the metal exposed regions provided at a plurality of locations on the plate surface are surrounded by the more hydrophobic hydrophobic surface region, so that the liquid sample is exposed to the metal.
  • the separated samples can be retained in the exposed metal region.
  • the liquid sample separated by chromatography can be stably held, and the analysis accuracy can be improved.
  • the Raman light is detected more efficiently than when a glass plate is used as the Raman analysis plate, for example. You can direct it to the vessel.
  • the sample is held in the exposed metal region, the background derived from glass generated when the glass plate is used can be suppressed, and the analysis accuracy can be further improved.
  • the exposed metal region is made of gold, silver, or aluminum.
  • the Raman analyzer according to the present invention is generated from the above-mentioned Raman analysis plate, a photoirradiator that irradiates the sample on the Raman analysis plate with light, and the sample by being irradiated with light. It is characterized by including a photodetector for detecting Raman light and an analysis unit for analyzing the sample based on the detected Raman spectrum. Since such a Raman analyzer is provided with the above-mentioned Raman analysis plate, it is possible to stably hold a liquid sample separated by chromatography, and it is possible to improve the analysis accuracy.
  • the liquid sample separated on the Raman analysis plate is waited to dry and the dried sample is irradiated with light for analysis.
  • Raman is used. Since the measurement range in the analysis is determined visually and it is difficult to visually find the dried sample, it takes time to determine the measurement range.
  • an image pickup device for imaging the surface of the Raman analysis plate is further provided, and the analysis unit includes the sample dried on the Raman analysis plate based on the image pickup data obtained by the image pickup device. It is preferable to determine the measurement range and analyze the measurement range. With such a configuration, the measurement range can be determined in a short time without relying on visual inspection.
  • One of the present inventions is an analysis system including the above-mentioned Raman analyzer, a separation column in which the liquid sample is sent by a mobile phase, and a separation mechanism for separating the sample by chromatography.
  • the liquid sample separated by chromatography can be stably held. Further, by drying the separated liquid sample on a Raman analysis plate, the sample is concentrated, so that the detection sensitivity of the small molecule compound contained in the sample can be improved to 1 ⁇ M or less, for example.
  • the analysis unit analyzes the sample based on a plurality of Raman spectra obtained by changing the mobile phase.
  • a peak with no fluctuation or little fluctuation can be used as a sample. It can be regarded as derived.
  • the spectrum derived from the sample can be isolated from the obtained Raman spectrum, and the analysis accuracy can be improved.
  • the separation mechanism separates a fluorescent substance that emits light in a wavelength band that overlaps with at least a part of the Raman light wavelength band generated from the sample from the sample.
  • light having an excitation wavelength of a fluorescent substance can be used as the light to irradiate the sample, which contributes to various analyzes according to various purposes and the like.
  • the Raman analysis method according to the present invention is a Raman analysis method in which a liquid sample separated by chromatography is held on a Raman analysis plate and analyzed by Raman spectroscopy, and the Raman analysis plate is a plate. It is provided with a metal exposed region provided at a plurality of locations on the surface to hold the sample, and a hydrophobic surface region provided on the plate surface and surrounding the metal exposed region and having a higher hydrophobicity than the metal exposed region. It is a method characterized by. According to such a Raman analysis method, a liquid sample separated by chromatography can be stably held, similar to the action and effect of the Raman analysis plate described above.
  • the liquid sample separated by chromatography can be stably held on the Raman analysis plate, and the analysis accuracy of Raman analysis can be improved.
  • the schematic diagram which shows the whole structure of the analysis system in one Embodiment of this invention The schematic diagram which shows the structure of the Raman analysis plate of the same embodiment.
  • the analysis system 100 of the present embodiment is a system that utilizes both chromatography and Raman spectroscopy, and specifically, as shown in FIG. 1, a separation mechanism 10 that separates a liquid sample S by liquid chromatography. And a Raman analyzer 20 that analyzes the separated sample S by Raman spectroscopy.
  • the separation mechanism 10 may be a mechanism that separates the liquid sample S by supercritical fluid chromatography.
  • the separation mechanism 10 sucks the mobile phase Z stored in the storage unit 11 into the flow path 13 by the pump 12, injects the liquid sample S into the flow path 13, and together with the mobile phase Z.
  • the sample S is configured to be separated for each component.
  • the mobile phase Z is, for example, a mixed liquid in which a plurality of types of liquids are mixed, and here, a mixed liquid of water and an organic solvent such as ethanol.
  • the mobile phase Z may be a single liquid or a gradient solvent having a concentration gradient.
  • the Raman analyzer 20 includes a Raman analysis plate 21 that holds the sample S, a light irradiator 22 that irradiates the sample S on the Raman analysis plate 21 with excitation light such as laser light, and the like.
  • a spectroscope 23 that disperses the Raman scattered light generated from the sample S by being irradiated with the excitation light, a light detector 24 that detects the dispersed Raman scattered light, and the sample S based on the detected Raman spectrum. It is provided with an analysis unit 25 for analysis. A specific embodiment of the analysis unit 25 will be described later.
  • the liquid sample S separated by the separation mechanism 10 is separated into a Raman analysis plate 21, and the sample S is dried and dried.
  • the sample S is configured to be analyzed by the Raman analyzer 20.
  • the Raman analysis plate 21 is used for analyzing the liquid sample S separated by chromatography by Raman spectroscopy, and the separated liquid sample S is dropped one after another. Is to be done.
  • the Raman analysis plate 21 here is, for example, a rectangular flat plate, but the shape is not limited to this, and the surface on which the liquid sample S is dropped, such as a disk or a block, is formed. If you have one, you may change it as appropriate.
  • the Raman analysis plate 21 is provided at a plurality of locations on the plate surface to surround the metal exposed region 211 for holding the sample S and the metal exposed region 211 provided on the plate surface. It is provided with a hydrophobic region 212 having a higher hydrophobicity than the metal exposed region 211.
  • the plate surface referred to here is shown in the upper part of FIG. 2, and is a surface that can be visually recognized from the outside of the Raman analysis plate 21, in other words, an exposed surface of the Raman analysis plate 21.
  • a plurality of exposed metal regions 211 and hydrophobic regions 212 surrounding each of the exposed metal regions 211 are provided on one side of a flat plate-shaped plate body 213 made of glass or the like.
  • a liquid-repellent film 214 is first formed on one side of the plate body 213, and for example, a metal is spot-welded to the surface of the liquid-repellent film 214.
  • the surface of the metal spot 215 is formed as the metal exposed region 211
  • the exposed portion of the liquid repellent film 214 is formed as the hydrophobic region 212 around the metal spot 215.
  • the exposed metal region 211 is made of a metal having high reflectance such as gold, silver, or aluminum, and has a circular shape having a diameter of, for example, about 100 ⁇ m to 300 ⁇ m.
  • the plurality of exposed metal regions 211 are arranged at equal intervals along the vertical direction on one side of the plate body 213 and at equal intervals along the horizontal direction.
  • the hydrophobic region 212 is made of a liquid-repellent film 214 formed by, for example, chemical modification or a periodic microstructure, and is provided here in all regions except the metal exposed region 211 on one side of the plate body 213. There is. However, the hydrophobic region 212 may be provided so as to surround each of the metal exposed regions 211, and does not necessarily have to be provided in all regions except the metal exposed region 211.
  • the liquid sample S separated on the Raman analysis plate 21 is dried to dryness before analysis.
  • the sample S can be concentrated, and as a result, the Raman analyzer 20 in the analysis system 100 of the present embodiment can improve the detection sensitivity of the small molecule compound contained in the sample S to, for example, 1 ⁇ M or less. can.
  • the dried sample S is difficult to visually confirm, and when the measurement area by the Raman analyzer 20 is visually determined, it takes time to determine the measurement area.
  • the Raman analyzer 20 of the present embodiment further includes an image pickup device that images the surface of the Raman analysis plate 21, and the image pickup data obtained by this image pickup device is analyzed by the analysis unit 25.
  • the analysis unit 25 is configured to determine the measurement area based on the image pickup data.
  • the image pickup apparatus is configured to be movable in the vertical and horizontal directions along the plate surface with respect to the Raman analysis plate 21 or the Raman analysis plate 21 with respect to the image pickup apparatus. There is. Along with this movement, the image pickup apparatus sequentially captures images or moving images of each of the above-mentioned metal exposed regions 211, and sequentially outputs the image pickup data to the analysis unit 25.
  • the analysis unit 25 includes a CPU, a memory, and the like, and when each device cooperates according to the program stored in the memory, at least the measurement area determination unit 251 and the analysis execution unit 25 are as shown in FIG. It exerts a function as 252.
  • the measurement area determination unit 251 acquires the image pickup data obtained by the image pickup apparatus, identifies the sample S dried on the Raman analysis plate 21 by performing image analysis of the image pickup data, for example, and obtains the sample S.
  • the range to be included is determined as the measurement range.
  • the liquid sample S is held in the metal exposed region 211 of the Raman analysis plate 21 and dried, and the position of the dried sample S is, for example, a coffee ring effect. It can be mentioned how to use and identify. More specifically, for example, the sample S is discriminated from the sample S by comparing the brightness and the like in the bright field image with the threshold value, and the sample S is compared with the image shown by the imaging data and the reference image prepared in advance. There is a method of distinguishing between and other parts.
  • the analysis execution unit 252 sequentially performs Raman analysis for each measurement area determined by the measurement area determination unit 251. More specifically, when the measurement area determination unit 251 determines the measurement areas, the light from the light irradiator 22 is sequentially irradiated to those measurement areas, and the Raman scattered light generated from the sample S in each measurement area is generated. It is separated by the spectroscope 23 and detected by the light detector 24. Then, the analysis execution unit 252 analyzes, for example, the molecular structure and physical properties of the sample S in the measurement region based on the Raman spectrum detected from one measurement region, and performs this Raman analysis on the sample S in each measurement region. The analysis results are output to a display or the like.
  • the exposed metal regions 211 provided at a plurality of locations on the plate surface of the Raman analysis plate 21 are surrounded by a more hydrophobic surface region.
  • the separated sample S can be retained in the metal exposed region 211.
  • the liquid sample S separated by chromatography can be stably held, and the analysis accuracy can be improved.
  • the sample S is held in the metal exposed region 211 and the Raman light generated from the sample S is reflected in the metal exposed region 211, it is more efficient than the case where a glass plate is used as the Raman analysis plate 21. Raman light can be directed at the photodetector 24.
  • the measurement range in each metal exposed region 211 is determined based on the imaging data obtained by the imaging device, the measurement range can be determined in a short time without relying on visual inspection.
  • the present invention is not limited to the above embodiment.
  • the analysis unit 25 may be configured to analyze the sample S based on a plurality of Raman spectra obtained by changing the mobile phase Z.
  • the analysis unit 25 for example, a plurality of Ramans obtained when the ratios of a plurality of types of liquids (water and an organic solvent in the above embodiment) constituting the mobile phase Z are changed and the respective mobile phases Z are used.
  • An aspect of analyzing the sample S based on the spectrum can be mentioned. Specifically, among the obtained plurality of Raman spectra, for example, a peak having a large fluctuation can be regarded as being derived from the mobile phase Z, and a peak having no fluctuation or having a small fluctuation is derived from the sample S. Can be regarded. Thereby, the spectrum of the sample S can be isolated from the obtained Raman spectrum, and the analysis accuracy can be improved.
  • the composition ratio of liquid A and liquid B is, for example, 100: 0, 80:20, 40:60, 20:80. , 0: 100, and the Raman spectrum of the sample S is obtained using each solvent. Then, the Raman spectrum is measured by focusing on the peaks of the obtained Raman spectra that appear at the same time. At this time, it can be determined that the peak whose height fluctuates according to the composition ratio of the solvent is derived from the solvent, and conversely, the peak which does not fluctuate is derived from the sample.
  • each liquid is used alone as the mobile phase Z, and each liquid is used.
  • An aspect of analyzing the sample S based on a plurality of Raman spectra obtained when used can be mentioned. Specifically, a sample is obtained by subtracting the Raman spectrum obtained when the liquids are used alone as the mobile phase Z from the Raman spectrum obtained by using the mobile phase Z in which a plurality of types of liquids are mixed. The peak derived from S can be analyzed. Further, the peak derived from the sample S can also be analyzed by performing, for example, singular value decomposition or multivariate analysis of these plurality of spectra.
  • the analysis unit 25 may use a plurality of types of gradient solvents having different concentration gradients and analyze the sample S based on the Raman spectrum obtained when each gradient solvent is used.
  • a fluorescent substance that emits light in a wavelength band that overlaps with at least a part of the wavelength band of Raman light generated from the sample S is separated from the sample S. It may be.
  • the wavelength of fluorescence emitted in advance is known for the fluorescent substance to be separated (for example, an aromatic compound or a compound having a ring structure).
  • light having an excitation wavelength of a fluorescent substance for example, 480 nm to 550 nm
  • the fraction after separating the fluorescent substance is detected by a detector such as a UV / UV-VIS detector, a photodiode array detector (PDA), or a diode array (DAD). Detection may be performed to confirm that the fluorescent substance is separated.
  • a detector such as a UV / UV-VIS detector, a photodiode array detector (PDA), or a diode array (DAD). Detection may be performed to confirm that the fluorescent substance is separated.
  • the liquid repellent film 214 is formed on one side of the plate body 213, and the metal spot 215 is formed on the surface of the liquid repellent film 214, whereby the metal exposed region 211 and the hydrophobic region 212 are formed.
  • a metal film 216 is formed on one side of the plate body 213 by, for example, vapor deposition, and a liquid repellent film 214 is formed on the surface of the metal film 216 to form a metal exposed region.
  • the 211 and the hydrophobic region 212 may be formed.
  • the Raman analysis plate 21 of the above embodiment has a metal exposed region 211 and a hydrophobic region 212 provided on a plate body 213 such as glass, but if the plate body 213 is made of metal, for example.
  • a liquid-repellent film 214 By forming, for example, a liquid-repellent film 214 on a part of the surface thereof, a region of the plate body 213 where the liquid-repellent film 214 is not formed becomes a metal exposed region 211, and the surface of the liquid-repellent film 214 is a hydrophobic region. It becomes 212.
  • the glass plate which is the plate body 213 in the embodiment, can be eliminated.
  • the embodiment in which the liquid sample separated by chromatography is held on the Raman analysis plate 21 for analysis has been described, but the liquid sample before being separated by chromatography is held on the Raman analysis plate 21. It may be held at 21 for analysis.
  • a liquid sample separated by chromatography can be stably held.

Abstract

To provide a Raman analysis plate that is capable of stably holding a liquid sample separated through chromatography, in this invention, a Raman analysis plate 21 used for Raman spectroscopy of a liquid sample S separated through chromatography is made to comprise metal exposure areas 211 that are provided in a plurality of locations on the surface of the plate and hold the sample S and a hydrophobic area 212 that is provided on the surface of the plate, surrounds the metal exposure areas 211, and is more hydrophobic than the metal exposure areas 211.

Description

ラマン分析用プレート、ラマン分析装置、分析システム、及びラマン分析方法Raman analysis plate, Raman analyzer, analysis system, and Raman analysis method
　本発明は、ラマン分析用プレートやこれを用いたラマン分析装置などに関するものである。 The present invention relates to a Raman analysis plate and a Raman analyzer using the plate.
　ラマン分析装置としては、特許文献１に示すように、クロマトグラフィーにより分離した液状の試料をラマン分光法により分析するように構成されたものがある。 As a Raman analyzer, as shown in Patent Document 1, there is a device configured to analyze a liquid sample separated by chromatography by Raman spectroscopy.
　具体的にこの装置は、分離された液状の試料をガラスプレートや撥水性プレートの複数箇所に分取して乾固させ、その乾固した試料のラマンスペクトルを取得するものである。 Specifically, this device separates the separated liquid sample into multiple locations on a glass plate or water-repellent plate and dries it, and acquires the Raman spectrum of the dried sample.
　しかしながら、上述したようにガラスプレートや撥水性プレートを用いると、分取した液状の試料がプレート上で移動してしまい（例えば、試料が有機溶媒であれば濡れが生じて液滴が広がってしまい）、安定して保持することが難しい。 However, when a glass plate or a water-repellent plate is used as described above, the separated liquid sample moves on the plate (for example, if the sample is an organic solvent, wetting occurs and droplets spread. ), It is difficult to hold it stably.
国際公開２０１４／０２７６５２号International Publication 2014/027652
　そこで、本発明は、クロマトグラフィーにより分離された液状の試料を安定して保持することのできるラマン分析用プレートを提供することをその主たる課題とするものである。 Therefore, the main object of the present invention is to provide a plate for Raman analysis capable of stably holding a liquid sample separated by chromatography.
　すなわち本発明に係るラマン分析用プレートは、クロマトグラフィーにより分離された液状の試料をラマン分光法により分析するために用いられるラマン分析用プレートであって、プレート表面の複数箇所に設けられて、前記試料を保持する金属露出領域と、プレート表面に設けられて前記金属露出領域を取り囲み、前記金属露出領域よりも疎水性の高い疎水性領域とを備えることを特徴とするものである。 That is, the Raman analysis plate according to the present invention is a Raman analysis plate used for analyzing a liquid sample separated by chromatography by Raman spectroscopy, and is provided at a plurality of locations on the plate surface and described above. It is characterized by having a metal exposed region for holding a sample and a hydrophobic region provided on the surface of the plate and surrounding the metal exposed region and having a higher hydrophobicity than the metal exposed region.
　このように構成されたラマン分析用プレートによれば、プレート表面の複数箇所に設けられた金属露出領域が、より疎水性の高い疎水性表領域に取り囲まれているので、液状の試料を金属露出領域に分取することで、分取した試料を金属露出領域に留まらせることができる。これにより、クロマトグラフィーにより分離された液状の試料を安定して保持することができ、ひいては分析精度の向上を図れる。
　さらに、金属露出領域に試料を保持させており、試料から発生したラマン光が金属露出領域で反射するので、ラマン分析用プレートとして例えばガラスプレートを用いる場合に比べて、効率良くラマン光を光検出器に向かわせることができる。そのうえ、金属露出領域に試料を保持させているので、ガラスプレートを用いた場合に生じるガラス由来のバックグラウンドを抑えることができ、分析精度のさらなる向上を図れる。 According to the Raman analysis plate configured in this way, the metal exposed regions provided at a plurality of locations on the plate surface are surrounded by the more hydrophobic hydrophobic surface region, so that the liquid sample is exposed to the metal. By separating into regions, the separated samples can be retained in the exposed metal region. As a result, the liquid sample separated by chromatography can be stably held, and the analysis accuracy can be improved.
Furthermore, since the sample is held in the exposed metal region and the Raman light generated from the sample is reflected in the exposed metal region, the Raman light is detected more efficiently than when a glass plate is used as the Raman analysis plate, for example. You can direct it to the vessel. Moreover, since the sample is held in the exposed metal region, the background derived from glass generated when the glass plate is used can be suppressed, and the analysis accuracy can be further improved.
　金属露出領域を反射率の高いものとするためには、前記金属露出領域が、金、銀、又はアルミニウムからなることが好ましい。 In order to make the exposed metal region highly reflective, it is preferable that the exposed metal region is made of gold, silver, or aluminum.
　また、本発明に係るラマン分析装置は、上述したラマン分析用プレートと、前記ラマン分析用プレート上の前記試料に光を照射する光照射器と、光が照射されることにより前記試料から発生するラマン光を検出する光検出器と、検出されたラマンスペクトルに基づいて前記試料を分析する分析部とを備えることを特徴とするものである。
　このようなラマン分析装置であれば、上述したラマン分析用プレートを備えているので、クロマトグラフィーにより分離された液状の試料を安定して保持することができ、ひいては分析精度の向上を図れる。 Further, the Raman analyzer according to the present invention is generated from the above-mentioned Raman analysis plate, a photoirradiator that irradiates the sample on the Raman analysis plate with light, and the sample by being irradiated with light. It is characterized by including a photodetector for detecting Raman light and an analysis unit for analyzing the sample based on the detected Raman spectrum.
Since such a Raman analyzer is provided with the above-mentioned Raman analysis plate, it is possible to stably hold a liquid sample separated by chromatography, and it is possible to improve the analysis accuracy.
　上述したラマン分析装置を用いる場合、ラマン分析用プレートに分取した液状の試料が乾固するのを待ち、乾固した試料に光を照射して分析することになるが、一般的に、ラマン分析における測定範囲の決定は目視により行われており、乾固した試料は目視では見つけ難いことから、測定範囲を決定するまでに時間を要する。 When the above-mentioned Raman analyzer is used, the liquid sample separated on the Raman analysis plate is waited to dry and the dried sample is irradiated with light for analysis. Generally, Raman is used. Since the measurement range in the analysis is determined visually and it is difficult to visually find the dried sample, it takes time to determine the measurement range.
　そこで、前記ラマン分析用プレートの表面を撮像する撮像装置をさらに備え、前記分析部が、前記撮像装置により得られた撮像データに基づいて、前記ラマン分析用プレート上で乾固した前記試料を含む測定範囲を決定し、その測定範囲を分析することが好ましい。
　このような構成であれば、目視に頼ることなく、短時間で測定範囲を決定することができる。 Therefore, an image pickup device for imaging the surface of the Raman analysis plate is further provided, and the analysis unit includes the sample dried on the Raman analysis plate based on the image pickup data obtained by the image pickup device. It is preferable to determine the measurement range and analyze the measurement range.
With such a configuration, the measurement range can be determined in a short time without relying on visual inspection.
　上述したラマン分析装置と、移動相により液状の前記試料が送液される分離カラムを備え、前記試料をクロマトグラフィーにより分離する分離機構とを具備する分析システムも本発明の１つである。
　このような分析システムによれば、上述したラマン分析用プレートを備えているので、クロマトグラフィーにより分離した液状の試料を安定して保持することができる。
　さらに、分離した液状の試料をラマン分析用プレート上で乾固させることにより、試料が濃縮するので、例えば試料に含まれる低分子化合物の検出感度を１μＭ以下まで向上させることができる。 One of the present inventions is an analysis system including the above-mentioned Raman analyzer, a separation column in which the liquid sample is sent by a mobile phase, and a separation mechanism for separating the sample by chromatography.
According to such an analysis system, since the above-mentioned Raman analysis plate is provided, the liquid sample separated by chromatography can be stably held.
Further, by drying the separated liquid sample on a Raman analysis plate, the sample is concentrated, so that the detection sensitivity of the small molecule compound contained in the sample can be improved to 1 μM or less, for example.
　ところで、移動相に由来する信号と試料に由来する信号とが重なり合うと、試料由来の信号が弱い場合に埋もれてしまい、精度良く分析することができなくなる。
　そこで、前記分析部が、前記移動相を変えて得られる複数のラマンスペクトルに基づいて、前記試料を分析することが好ましい。
　このような構成であれば、例えば移動相の濃度を変えて複数回測定し、それら複数回の測定により得られたラマンスペクトルを比較することで、例えば変動の無い或いは変動の小さいピークを試料に由来するものとみなすことができる。これにより、得られたラマンスペクトルから試料由来のスペクトルを単離することができ、分析精度の向上を図れる。 By the way, when the signal derived from the mobile phase and the signal derived from the sample overlap, the signal derived from the sample is buried when the signal is weak, and accurate analysis cannot be performed.
Therefore, it is preferable that the analysis unit analyzes the sample based on a plurality of Raman spectra obtained by changing the mobile phase.
With such a configuration, for example, by changing the concentration of the mobile phase and measuring multiple times and comparing the Raman spectra obtained by these multiple measurements, for example, a peak with no fluctuation or little fluctuation can be used as a sample. It can be regarded as derived. As a result, the spectrum derived from the sample can be isolated from the obtained Raman spectrum, and the analysis accuracy can be improved.
　前記分離機構が、前記試料から発生するラマン光の波長帯の少なくとも一部と重なる波長帯の光を発する蛍光物質を前記試料から分離することが好ましい。
　このような構成であれば、試料に照射する光として蛍光物質の励起波長の光を用いることができ、様々な目的等に応じた種々の分析に資する。 It is preferable that the separation mechanism separates a fluorescent substance that emits light in a wavelength band that overlaps with at least a part of the Raman light wavelength band generated from the sample from the sample.
With such a configuration, light having an excitation wavelength of a fluorescent substance can be used as the light to irradiate the sample, which contributes to various analyzes according to various purposes and the like.
　また、本発明に係るラマン分析方法は、クロマトグラフィーにより分離された液状の試料をラマン分析用プレートに保持させてラマン分光法により分析するラマン分析方法であって、前記ラマン分析用プレートが、プレート表面の複数箇所に設けられて、前記試料を保持する金属露出領域と、プレート表面に設けられて前記金属露出領域を取り囲み、前記金属露出領域よりも疎水性の高い疎水性表領域とを備えることを特徴とする方法である。
　このようなラマン分析方法によれば、上述したラマン分析用プレートによる作用効果と同様に、クロマトグラフィーにより分離された液状の試料を安定して保持することができる。 Further, the Raman analysis method according to the present invention is a Raman analysis method in which a liquid sample separated by chromatography is held on a Raman analysis plate and analyzed by Raman spectroscopy, and the Raman analysis plate is a plate. It is provided with a metal exposed region provided at a plurality of locations on the surface to hold the sample, and a hydrophobic surface region provided on the plate surface and surrounding the metal exposed region and having a higher hydrophobicity than the metal exposed region. It is a method characterized by.
According to such a Raman analysis method, a liquid sample separated by chromatography can be stably held, similar to the action and effect of the Raman analysis plate described above.
　このように構成した本発明によれば、クロマトグラフィーにより分離された液状の試料をラマン分析用プレート上で安定して保持することができ、ひいてはラマン分析の分析精度を向上させることができる。 According to the present invention configured as described above, the liquid sample separated by chromatography can be stably held on the Raman analysis plate, and the analysis accuracy of Raman analysis can be improved.
本発明の一実施形態における分析システムの全体構成を示す模式図。The schematic diagram which shows the whole structure of the analysis system in one Embodiment of this invention. 同実施形態のラマン分析用プレートの構成を示す模式図。The schematic diagram which shows the structure of the Raman analysis plate of the same embodiment. 同実施形態のラマン分析装置を構成する撮像装置を説明するための模式図。The schematic diagram for demonstrating the image pickup apparatus which comprises the Raman analysis apparatus of the same embodiment. 同実施形態の分析部の機能を示す機能ブロック図。The functional block diagram which shows the function of the analysis part of the same embodiment. その他の実施形態の分析部の態様を説明ための図。The figure for demonstrating the aspect of the analysis part of another embodiment. その他の実施形態のラマン分析用プレートの構成を示す模式図。The schematic diagram which shows the structure of the Raman analysis plate of another embodiment.
１００・・・分析システム
Ｓ　　・・・試料
Ｚ　　・・・移動相
１０　・・・分離機構
２０　・・・ラマン分析装置
２１　・・・ラマン分析用プレート
２１１・・・金属露出領域
２１２・・・疎水性領域
２１３・・・プレート本体
２１４・・・撥液膜
２１５・・・金属スポット
２１６・・・金属膜 100 ... Analytical system S ... Sample Z ... Mobile phase 10 ... Separation mechanism 20 ... Raman analyzer 21 ... Raman analysis plate 211 ... Metal exposed area 212 ... Hydrophobic Sexual region 213 ... Plate body 214 ... Liquid repellent film 215 ... Metal spot 216 ... Metal film
　以下に本発明に係る分析システムの一実施形態について図面を参照して説明する。 Hereinafter, an embodiment of the analysis system according to the present invention will be described with reference to the drawings.
　本実施形態の分析システム１００は、クロマトグラフィーとラマン分光法との双方を利用したシステムであり、具体的には図１に示すように、液体クロマトグラフィーにより液状の試料Ｓを分離する分離機構１０と、分離された試料Ｓをラマン分光法により分析するラマン分析装置２０とを備える。なお、分離機構１０としては、超臨界流体クロマトグラフィーにより液状の試料Ｓを分離するものであっても良い。 The analysis system 100 of the present embodiment is a system that utilizes both chromatography and Raman spectroscopy, and specifically, as shown in FIG. 1, a separation mechanism 10 that separates a liquid sample S by liquid chromatography. And a Raman analyzer 20 that analyzes the separated sample S by Raman spectroscopy. The separation mechanism 10 may be a mechanism that separates the liquid sample S by supercritical fluid chromatography.
　分離機構１０は、図１に示すように、貯留部１１に貯留された移動相Ｚをポンプ１２により流路１３に吸い上げるとともに、その流路１３に液状の試料Ｓを注入し、移動相Ｚとともに試料Ｓを分離カラム１４に送液することで、試料Ｓを成分ごとに分離するように構成されたものである。
　なお、移動相Ｚは、例えば複数種類の液体が混合された混合液であり、ここでは水とエタノール等の有機溶媒との混合液である。ただし、移動相Ｚとしては、単一の液体からなるものであっても良いし、濃度勾配を有するグラジエント溶媒であっても良い。 As shown in FIG. 1, the separation mechanism 10 sucks the mobile phase Z stored in the storage unit 11 into the flow path 13 by the pump 12, injects the liquid sample S into the flow path 13, and together with the mobile phase Z. By sending the sample S to the separation column 14, the sample S is configured to be separated for each component.
The mobile phase Z is, for example, a mixed liquid in which a plurality of types of liquids are mixed, and here, a mixed liquid of water and an organic solvent such as ethanol. However, the mobile phase Z may be a single liquid or a gradient solvent having a concentration gradient.
　ラマン分析装置２０は、図１に示すように、試料Ｓを保持するラマン分析用プレート２１と、ラマン分析用プレート２１上の試料Ｓにレーザ光等の励起光を照射する光照射器２２と、励起光が照射されることにより試料Ｓから発生するラマン散乱光を分光する分光器２３と、分光されたラマン散乱光を検出する光検出器２４と、検出されたラマンスペクトルに基づいて試料Ｓを分析する分析部２５とを備える。なお、分析部２５の具体的な実施態様については後述する。 As shown in FIG. 1, the Raman analyzer 20 includes a Raman analysis plate 21 that holds the sample S, a light irradiator 22 that irradiates the sample S on the Raman analysis plate 21 with excitation light such as laser light, and the like. A spectroscope 23 that disperses the Raman scattered light generated from the sample S by being irradiated with the excitation light, a light detector 24 that detects the dispersed Raman scattered light, and the sample S based on the detected Raman spectrum. It is provided with an analysis unit 25 for analysis. A specific embodiment of the analysis unit 25 will be described later.
　然して、本実施形態の分析システム１００は、図１に示すように、分離機構１０により分離された液状の試料Ｓをラマン分析用プレート２１に分取し、その試料Ｓを乾固させ、乾固した試料Ｓをラマン分析装置２０により分析するように構成されている。 Therefore, in the analysis system 100 of the present embodiment, as shown in FIG. 1, the liquid sample S separated by the separation mechanism 10 is separated into a Raman analysis plate 21, and the sample S is dried and dried. The sample S is configured to be analyzed by the Raman analyzer 20.
　ラマン分析用プレート２１は、図２に示すように、クロマトグラフィーにより分離された液状の試料Ｓをラマン分光法により分析するために用いられるものであり、分離された液状の試料Ｓが次々と滴下されるものである。なお、ここでのラマン分析用プレート２１は、例えば矩形平板状のものであるが、形状はこれに限らず、例えば円板状やブロック状のものなど、液状の試料Ｓが滴下される表面を有するものであれば適宜変更して構わない。 As shown in FIG. 2, the Raman analysis plate 21 is used for analyzing the liquid sample S separated by chromatography by Raman spectroscopy, and the separated liquid sample S is dropped one after another. Is to be done. The Raman analysis plate 21 here is, for example, a rectangular flat plate, but the shape is not limited to this, and the surface on which the liquid sample S is dropped, such as a disk or a block, is formed. If you have one, you may change it as appropriate.
　そして、このラマン分析用プレート２１は、図２に示すように、プレート表面の複数箇所に設けられて、試料Ｓを保持する金属露出領域２１１と、プレート表面に設けられて金属露出領域２１１を取り囲み、金属露出領域２１１よりも疎水性の高い疎水性領域２１２とを備えるものである。なお、ここでいうプレート表面は、図２の上段に示されており、ラマン分析用プレート２１の外部から視認可能な面であって、言い換えればラマン分析用プレート２１の露出表面である。 Then, as shown in FIG. 2, the Raman analysis plate 21 is provided at a plurality of locations on the plate surface to surround the metal exposed region 211 for holding the sample S and the metal exposed region 211 provided on the plate surface. It is provided with a hydrophobic region 212 having a higher hydrophobicity than the metal exposed region 211. The plate surface referred to here is shown in the upper part of FIG. 2, and is a surface that can be visually recognized from the outside of the Raman analysis plate 21, in other words, an exposed surface of the Raman analysis plate 21.
　本実施形態では、ガラス等からなる平板状のプレート本体２１３の片面に複数の金属露出領域２１１と、それらの金属露出領域２１１それぞれを取り囲む疎水性領域２１２とが設けられている。 In the present embodiment, a plurality of exposed metal regions 211 and hydrophobic regions 212 surrounding each of the exposed metal regions 211 are provided on one side of a flat plate-shaped plate body 213 made of glass or the like.
　より具体的に説明すると、この実施形態では、まずプレート本体２１３の片面に撥液膜２１４を形成し、その撥液膜２１４の表面に例えば金属をスポット溶接してある。これにより、金属スポット２１５の表面が金属露出領域２１１として形成されるとともに、金属スポット２１５の周囲において撥液膜２１４の露出した部分が疎水性領域２１２として形成される。 More specifically, in this embodiment, a liquid-repellent film 214 is first formed on one side of the plate body 213, and for example, a metal is spot-welded to the surface of the liquid-repellent film 214. As a result, the surface of the metal spot 215 is formed as the metal exposed region 211, and the exposed portion of the liquid repellent film 214 is formed as the hydrophobic region 212 around the metal spot 215.
　金属露出領域２１１は、例えば金、銀、又はアルミニウムなどの反射率の高い金属からなるものであり、例えば直径１００μｍ～３００μｍ程度の円形状をなす。ここでは、複数の金属露出領域２１１が、プレート本体２１３の片面において縦方向に沿って等間隔に配置されるとともに、横方向に沿って等間隔に配置されている。 The exposed metal region 211 is made of a metal having high reflectance such as gold, silver, or aluminum, and has a circular shape having a diameter of, for example, about 100 μm to 300 μm. Here, the plurality of exposed metal regions 211 are arranged at equal intervals along the vertical direction on one side of the plate body 213 and at equal intervals along the horizontal direction.
　疎水性領域２１２は、例えば化学修飾や周期性微細構造などにより形成された撥液膜２１４からなるものであり、ここでは、プレート本体２１３の片面における金属露出領域２１１を除く全領域に設けられている。ただし、疎水性領域２１２としては、金属露出領域２１１それぞれの周囲を取り囲むように設けられていれば良く、必ずしも金属露出領域２１１を除く全領域に設けられている必要はない。 The hydrophobic region 212 is made of a liquid-repellent film 214 formed by, for example, chemical modification or a periodic microstructure, and is provided here in all regions except the metal exposed region 211 on one side of the plate body 213. There is. However, the hydrophobic region 212 may be provided so as to surround each of the metal exposed regions 211, and does not necessarily have to be provided in all regions except the metal exposed region 211.
　ところで、このラマン分析用プレート２１に分取した液状の試料Ｓは、上述したように、分析する前に乾固される。これにより試料Ｓを濃縮させることができ、その結果、本実施形態の分析システム１００におけるラマン分析装置２０であれば、例えば試料Ｓに含まれる低分子化合物の検出感度を１μＭ以下まで向上させることができる。 By the way, as described above, the liquid sample S separated on the Raman analysis plate 21 is dried to dryness before analysis. As a result, the sample S can be concentrated, and as a result, the Raman analyzer 20 in the analysis system 100 of the present embodiment can improve the detection sensitivity of the small molecule compound contained in the sample S to, for example, 1 μM or less. can.
　一方、乾固した試料Ｓは目視では確認しにくく、ラマン分析装置２０による測定領域を目視により決定しようとすると、その決定に時間がかかってしまう。 On the other hand, the dried sample S is difficult to visually confirm, and when the measurement area by the Raman analyzer 20 is visually determined, it takes time to determine the measurement area.
　そこで、本実施形態のラマン分析装置２０は、図３に示すように、ラマン分析用プレート２１の表面を撮像する撮像装置をさらに備えており、この撮像装置により得られた撮像データを分析部２５に出力し、分析部２５が撮像データに基づいて測定領域を決定するように構成されている。 Therefore, as shown in FIG. 3, the Raman analyzer 20 of the present embodiment further includes an image pickup device that images the surface of the Raman analysis plate 21, and the image pickup data obtained by this image pickup device is analyzed by the analysis unit 25. The analysis unit 25 is configured to determine the measurement area based on the image pickup data.
　より具体的に説明すると、撮像装置がラマン分析用プレート２１に対して、或いは、ラマン分析用プレート２１が撮像装置に対して、プレート表面に沿った縦方向及び横方向に移動可能に構成されている。この移動に伴い、撮像装置は、上述した金属露出領域２１１それぞれの画像又は動画を順次撮像し、それらの撮像データを分析部２５に逐次出力する。 More specifically, the image pickup apparatus is configured to be movable in the vertical and horizontal directions along the plate surface with respect to the Raman analysis plate 21 or the Raman analysis plate 21 with respect to the image pickup apparatus. There is. Along with this movement, the image pickup apparatus sequentially captures images or moving images of each of the above-mentioned metal exposed regions 211, and sequentially outputs the image pickup data to the analysis unit 25.
　分析部２５は、ＣＰＵ、メモリ等を備えたものであり、前記メモリに格納されたプログラムに従って各機器が協働することにより、図４に示すように、少なくとも測定領域決定部２５１及び分析実行部２５２としての機能を発揮するものである。 The analysis unit 25 includes a CPU, a memory, and the like, and when each device cooperates according to the program stored in the memory, at least the measurement area determination unit 251 and the analysis execution unit 25 are as shown in FIG. It exerts a function as 252.
　測定領域決定部２５１は、撮像装置により得られた撮像データを取得し、その撮像データを例えば画像解析することにより、ラマン分析用プレート２１上で乾固した試料Ｓを特定し、その試料Ｓを含む範囲を測定範囲として決定する。
　なお、試料Ｓの具体的な特定方法としては、液状の試料Ｓをラマン分析用プレート２１の金属露出領域２１１に保持させて乾固させ、この乾固した試料Ｓの位置を例えばコーヒーリング効果を利用して特定する方法を挙げることができる。より詳細には、例えば明視野画像における輝度等と閾値とを比較して試料Ｓとそれ以外の箇所とを判別したり、撮像データが示す画像と予め準備した基準画像とを比較して試料Ｓとそれ以外の箇所とを判別したりする方法を挙げることができる。 The measurement area determination unit 251 acquires the image pickup data obtained by the image pickup apparatus, identifies the sample S dried on the Raman analysis plate 21 by performing image analysis of the image pickup data, for example, and obtains the sample S. The range to be included is determined as the measurement range.
As a specific method for specifying the sample S, the liquid sample S is held in the metal exposed region 211 of the Raman analysis plate 21 and dried, and the position of the dried sample S is, for example, a coffee ring effect. It can be mentioned how to use and identify. More specifically, for example, the sample S is discriminated from the sample S by comparing the brightness and the like in the bright field image with the threshold value, and the sample S is compared with the image shown by the imaging data and the reference image prepared in advance. There is a method of distinguishing between and other parts.
　分析実行部２５２は、測定領域決定部２５１が決定した測定領域それぞれに対して順次ラマン分析を行うものである。
　より具体的に説明すると、測定領域決定部２５１が測定領域を決定すると、それらの測定領域に光照射器２２からの光が順次照射され、それぞれの測定領域における試料Ｓから発生するラマン散乱光が分光器２３により分光されて光検出器２４により検出される。
　そして、分析実行部２５２は、１つの測定領域から検出されたラマンスペクトルに基づいて、その測定領域における試料Ｓの例えば分子構造や物性などを分析し、このラマン分析をそれぞれの測定領域の試料Ｓに対して順次行い、それらの分析結果をディスプレイ等に出力する。 The analysis execution unit 252 sequentially performs Raman analysis for each measurement area determined by the measurement area determination unit 251.
More specifically, when the measurement area determination unit 251 determines the measurement areas, the light from the light irradiator 22 is sequentially irradiated to those measurement areas, and the Raman scattered light generated from the sample S in each measurement area is generated. It is separated by the spectroscope 23 and detected by the light detector 24.
Then, the analysis execution unit 252 analyzes, for example, the molecular structure and physical properties of the sample S in the measurement region based on the Raman spectrum detected from one measurement region, and performs this Raman analysis on the sample S in each measurement region. The analysis results are output to a display or the like.
　このように構成された分析システム１００によれば、ラマン分析用プレート２１のプレート表面の複数箇所に設けられた金属露出領域２１１が、より疎水性の高い疎水性表領域に取り囲まれているので、液状の試料Ｓを金属露出領域２１１に分取することで、分取した試料Ｓを金属露出領域２１１に留まらせることができる。これにより、クロマトグラフィーにより分離された液状の試料Ｓを安定して保持することができ、ひいては分析精度の向上を図れる。 According to the analysis system 100 configured in this way, the exposed metal regions 211 provided at a plurality of locations on the plate surface of the Raman analysis plate 21 are surrounded by a more hydrophobic surface region. By separating the liquid sample S into the metal exposed region 211, the separated sample S can be retained in the metal exposed region 211. As a result, the liquid sample S separated by chromatography can be stably held, and the analysis accuracy can be improved.
　さらに、金属露出領域２１１に試料Ｓを保持させており、試料Ｓから発生したラマン光が金属露出領域２１１で反射するので、ラマン分析用プレート２１として例えばガラスプレートを用いる場合に比べて、効率良くラマン光を光検出器２４に向かわせることができる。 Further, since the sample S is held in the metal exposed region 211 and the Raman light generated from the sample S is reflected in the metal exposed region 211, it is more efficient than the case where a glass plate is used as the Raman analysis plate 21. Raman light can be directed at the photodetector 24.
　そのうえ、それぞれの金属露出領域２１１における測定範囲を、撮像装置により得られた撮像データに基づいて決定するので、測定範囲の決定を目視に頼ることなく短時間で行うことができる。 Moreover, since the measurement range in each metal exposed region 211 is determined based on the imaging data obtained by the imaging device, the measurement range can be determined in a short time without relying on visual inspection.
　なお、本発明は前記実施形態に限られるものではない。 The present invention is not limited to the above embodiment.
　例えば、移動相Ｚに由来する信号と試料Ｓに由来する信号とが重なり合うと、試料Ｓに由来の信号が弱い場合に埋もれてしまい、精度良く分析することができなくなる場合がある。
　そこで、分析部２５としては、図５に示すように、前記移動相Ｚを変えて得られる複数のラマンスペクトルに基づいて、試料Ｓを分析するように構成されていても良い。 For example, if the signal derived from the mobile phase Z and the signal derived from the sample S overlap, the signal derived from the sample S may be buried when the signal is weak, and accurate analysis may not be possible.
Therefore, as shown in FIG. 5, the analysis unit 25 may be configured to analyze the sample S based on a plurality of Raman spectra obtained by changing the mobile phase Z.
　かかる分析部２５の一例としては、例えば移動相Ｚを構成する複数種類の液体（前記実施形態では水と有機溶媒）の比率を変え、それぞれの移動相Ｚを用いた場合に得られる複数のラマンスペクトルに基づいて試料Ｓを分析する態様を挙げることができる。
　具体的には、得られた複数のラマンスペクトルのうち、例えば変動の大きいピークは移動相Ｚに由来するものとみなすことができ、変動の無い或いは変動の小さいピークを試料Ｓに由来するものとみなすことができる。これにより、得られたラマンスペクトルから試料Ｓのスペクトルを単離することができ、分析精度の向上を図れる。 As an example of the analysis unit 25, for example, a plurality of Ramans obtained when the ratios of a plurality of types of liquids (water and an organic solvent in the above embodiment) constituting the mobile phase Z are changed and the respective mobile phases Z are used. An aspect of analyzing the sample S based on the spectrum can be mentioned.
Specifically, among the obtained plurality of Raman spectra, for example, a peak having a large fluctuation can be regarded as being derived from the mobile phase Z, and a peak having no fluctuation or having a small fluctuation is derived from the sample S. Can be regarded. Thereby, the spectrum of the sample S can be isolated from the obtained Raman spectrum, and the analysis accuracy can be improved.
　より具体的な一例を挙げると、例えば溶媒が液体Ａと液体Ｂとから構成されている場合、液体Ａと液体Ｂの構成比率を例えば１００：０、８０：２０、４０：６０、２０：８０、０：１００にして、それぞれの溶媒を用いて試料Ｓのラマンスペクトルを取得する。
　そして、得られたそれぞれのラマンスペクトルの同じ時間に出たピークに着目してラマンスペクトルを測定する。
　この際、溶媒の構成比率に応じて高さが変動するピークは溶媒由来のものであり、逆に変動していないピークは試料由来のものと判断することができる。 To give a more specific example, for example, when the solvent is composed of liquid A and liquid B, the composition ratio of liquid A and liquid B is, for example, 100: 0, 80:20, 40:60, 20:80. , 0: 100, and the Raman spectrum of the sample S is obtained using each solvent.
Then, the Raman spectrum is measured by focusing on the peaks of the obtained Raman spectra that appear at the same time.
At this time, it can be determined that the peak whose height fluctuates according to the composition ratio of the solvent is derived from the solvent, and conversely, the peak which does not fluctuate is derived from the sample.
　また、複数のラマンスペクトルを用いる分析態様の別の一例としては、移動相Ｚが複数種類の液体を混合したものである場合、それぞれの液体を単体で移動相Ｚとして用いて、それぞれの液体を用いた場合に得られる複数のラマンスペクトルに基づいて試料Ｓを分析する態様を挙げることができる。
　具体的には、複数種類の液体を混合した移動相Ｚを用いて得られたラマンスペクトルから、それらの液体を単体で移動相Ｚとして用いた場合に得られたラマンスペクトルを差し引くことで、試料Ｓに由来するピークを解析することができる。
　また、これら複数のスペクトルを例えば特異値分解や多変量解析などすることによっても、試料Ｓに由来するピークを解析することができる。 Further, as another example of the analysis mode using a plurality of Raman spectra, when the mobile phase Z is a mixture of a plurality of types of liquids, each liquid is used alone as the mobile phase Z, and each liquid is used. An aspect of analyzing the sample S based on a plurality of Raman spectra obtained when used can be mentioned.
Specifically, a sample is obtained by subtracting the Raman spectrum obtained when the liquids are used alone as the mobile phase Z from the Raman spectrum obtained by using the mobile phase Z in which a plurality of types of liquids are mixed. The peak derived from S can be analyzed.
Further, the peak derived from the sample S can also be analyzed by performing, for example, singular value decomposition or multivariate analysis of these plurality of spectra.
　さらに分析部２５としては、濃度勾配の異なる複数種類のグラジエント溶媒を用いて、それぞれのグラジエント溶媒を用いた場合に得られるラマンスペクトルに基づいて試料Ｓを分析しても良い。 Further, the analysis unit 25 may use a plurality of types of gradient solvents having different concentration gradients and analyze the sample S based on the Raman spectrum obtained when each gradient solvent is used.
　分離機構１０としては、試料Ｓが自家蛍光を発しないものである場合、試料Ｓから発生するラマン光の波長帯の少なくとも一部と重なる波長帯の光を発する蛍光物質を試料Ｓから分離するものであっても良い。なお、分離対象となる蛍光物質（例えば、芳香族や環構造の化合物等）は予め発光する蛍光の波長が既知である。
　このような構成であれば、試料Ｓに照射する光として蛍光物質の励起波長（例えば４８０ｎｍ～５５０ｎｍ）の光を用いることができ、様々な目的等に応じた種々の分析に資する。なお、試料Ｓの測定前に、蛍光物質を分離した後の分画をUV/UV-VIS検出器、フォトダイオードアレイ検出器（PDA）、ダイオードアレイ（DAD）等の検出器にて蛍光物質の検出を行い、蛍光物質が分離されていることを確認しても良い。 As the separation mechanism 10, when the sample S does not emit autofluorescence, a fluorescent substance that emits light in a wavelength band that overlaps with at least a part of the wavelength band of Raman light generated from the sample S is separated from the sample S. It may be. The wavelength of fluorescence emitted in advance is known for the fluorescent substance to be separated (for example, an aromatic compound or a compound having a ring structure).
With such a configuration, light having an excitation wavelength of a fluorescent substance (for example, 480 nm to 550 nm) can be used as the light to irradiate the sample S, which contributes to various analyzes according to various purposes and the like. Before the measurement of the sample S, the fraction after separating the fluorescent substance is detected by a detector such as a UV / UV-VIS detector, a photodiode array detector (PDA), or a diode array (DAD). Detection may be performed to confirm that the fluorescent substance is separated.
　前記実施形態のラマン分析用プレート２１は、プレート本体２１３の片面に撥液膜２１４を形成し、その撥液膜２１４の表面に金属スポット２１５を形成することで金属露出領域２１１及び疎水性領域２１２を形成していたが、図６に示すように、プレート本体２１３の片面に金属膜２１６を例えば蒸着により形成し、その金属膜２１６の表面に撥液膜２１４を形成することで、金属露出領域２１１及び疎水性領域２１２を形成しても良い。 In the Raman analysis plate 21 of the above embodiment, the liquid repellent film 214 is formed on one side of the plate body 213, and the metal spot 215 is formed on the surface of the liquid repellent film 214, whereby the metal exposed region 211 and the hydrophobic region 212 are formed. However, as shown in FIG. 6, a metal film 216 is formed on one side of the plate body 213 by, for example, vapor deposition, and a liquid repellent film 214 is formed on the surface of the metal film 216 to form a metal exposed region. The 211 and the hydrophobic region 212 may be formed.
　また、前記実施形態のラマン分析用プレート２１は、ガラス等のプレート本体２１３に金属露出領域２１１及び疎水性領域２１２を設けたものであったが、例えばプレート本体２１３が金属製のものであれば、その表面の一部に例えば撥液膜２１４を生成することで、プレート本体２１３のうち撥液膜２１４が生成されていない領域が金属露出領域２１１となり、撥液膜２１４の表面が疎水性領域２１２となる。このような構成であれば、前記実施形態におけるプレート本体２１３であるガラスプレートを不要にすることができる。 Further, the Raman analysis plate 21 of the above embodiment has a metal exposed region 211 and a hydrophobic region 212 provided on a plate body 213 such as glass, but if the plate body 213 is made of metal, for example. By forming, for example, a liquid-repellent film 214 on a part of the surface thereof, a region of the plate body 213 where the liquid-repellent film 214 is not formed becomes a metal exposed region 211, and the surface of the liquid-repellent film 214 is a hydrophobic region. It becomes 212. With such a configuration, the glass plate, which is the plate body 213 in the embodiment, can be eliminated.
　なお、前記実施形態では、クロマトグラフィーにより分離された液状の試料をラマン分析用プレート２１に保持させて分析する態様について説明したが、クロマトグラフィーにより分離される前の液状の試料をラマン分析用プレート２１に保持させて分析しても良い。 In the above embodiment, the embodiment in which the liquid sample separated by chromatography is held on the Raman analysis plate 21 for analysis has been described, but the liquid sample before being separated by chromatography is held on the Raman analysis plate 21. It may be held at 21 for analysis.
　その他、本発明は前記実施形態に限られず、その趣旨を逸脱しない範囲で種々の変形が可能であるのは言うまでもない。 In addition, the present invention is not limited to the above-described embodiment, and it goes without saying that various modifications can be made without departing from the spirit of the present invention.
　本発明によれば、クロマトグラフィーにより分離された液状の試料を安定して保持することができる。
  According to the present invention, a liquid sample separated by chromatography can be stably held.

Claims (8)

  1. 　クロマトグラフィーにより分離された液状の試料をラマン分光法により分析するために用いられるラマン分析用プレートであって、
    　プレート表面の複数箇所に設けられて、前記試料を保持する金属露出領域と、
    　プレート表面に設けられて前記金属露出領域を取り囲み、前記金属露出領域よりも疎水性の高い疎水性領域とを備えることを特徴とするラマン分析用プレート。     A plate for Raman analysis used for analyzing a liquid sample separated by chromatography by Raman spectroscopy.
    Metal exposed areas provided at multiple locations on the plate surface to hold the sample,
    A plate for Raman analysis, which is provided on the surface of a plate, surrounds the exposed metal region, and has a hydrophobic region having a higher hydrophobicity than the exposed metal region.
  2. 　前記金属露出領域が、金、銀、又はアルミニウムからなることを特徴とする請求項１記載のラマン分析用プレート。 The Raman analysis plate according to claim 1, wherein the exposed metal region is made of gold, silver, or aluminum.
  3. 　請求項１又は２記載のラマン分析用プレートと、
    　前記ラマン分析用プレート上の前記試料に光を照射する光照射器と、
    　光が照射されることにより前記試料から発生するラマン光を検出する光検出器と、
    　検出されたラマンスペクトルに基づいて前記試料を分析する分析部とを備える、ラマン分析装置。     The Raman analysis plate according to claim 1 or 2,
    A light irradiator that irradiates the sample on the Raman analysis plate with light,
    A photodetector that detects Raman light generated from the sample when irradiated with light, and a photodetector.
    A Raman analyzer comprising an analysis unit that analyzes the sample based on the detected Raman spectrum.
  4. 　前記ラマン分析用プレートの表面を撮像する撮像装置をさらに備え、
    　前記分析部が、前記撮像装置により得られた撮像データに基づいて、前記ラマン分析用プレート上で乾固した前記試料を含む測定範囲を決定し、その測定範囲を分析する、請求項３記載のラマン分析装置。     An image pickup device for imaging the surface of the Raman analysis plate is further provided.
    The third aspect of the present invention, wherein the analysis unit determines a measurement range including the sample dried on the Raman analysis plate based on the image pickup data obtained by the image pickup apparatus, and analyzes the measurement range. Raman analyzer.
  5. 　請求項３又は４記載のラマン分析装置と、
    　移動相により液状の前記試料が送液される分離カラムを備え、当該試料をクロマトグラフィーにより分離する分離機構とを具備する、分析システム。     The Raman analyzer according to claim 3 or 4, and the Raman analyzer.
    An analytical system comprising a separation column through which the liquid sample is fed by a mobile phase and a separation mechanism for separating the sample by chromatography.
  6. 　前記分析部が、前記移動相を変えて得られる複数のラマンスペクトルに基づいて、前記試料を分析する、請求項５記載の分析システム。 The analysis system according to claim 5, wherein the analysis unit analyzes the sample based on a plurality of Raman spectra obtained by changing the mobile phase.
  7. 　前記分離機構が、前記試料から発生するラマン光の波長帯の少なくとも一部と重なる波長帯の光を発する蛍光物質を前記試料から分離する、請求項５又は６記載の分析システム。 The analysis system according to claim 5 or 6, wherein the separation mechanism separates a fluorescent substance that emits light in a wavelength band that overlaps at least a part of the Raman light wavelength band generated from the sample from the sample.
  8. 　クロマトグラフィーにより分離された液状の試料をラマン分析用プレートに保持させてラマン分光法により分析するラマン分析方法であって、
    　前記ラマン分析用プレートが、
    　プレート表面の複数箇所に設けられて、前記試料を保持する金属露出領域と、
    　プレート表面に設けられて前記金属露出領域を取り囲み、前記金属露出領域よりも疎水性の高い疎水性領域とを備えることを特徴とするラマン分析方法。
          It is a Raman analysis method in which a liquid sample separated by chromatography is held on a plate for Raman analysis and analyzed by Raman spectroscopy.
    The Raman analysis plate
    Metal exposed areas provided at multiple locations on the plate surface to hold the sample,
    A Raman analysis method provided on a plate surface, surrounding the exposed metal region, and having a hydrophobic region having a higher hydrophobicity than the exposed metal region.
PCT/JP2021/040560 2020-12-07 2021-11-04 Raman analysis plate, raman analysis device, analysis system, and raman analysis method WO2022123968A1 (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP2022568109A JPWO2022123968A1 (en) 2020-12-07 2021-11-04

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2020202452 2020-12-07
JP2020-202452 2020-12-07

Publications (1)

Publication Number Publication Date
WO2022123968A1 true WO2022123968A1 (en) 2022-06-16

Family

ID=81973640

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2021/040560 WO2022123968A1 (en) 2020-12-07 2021-11-04 Raman analysis plate, raman analysis device, analysis system, and raman analysis method

Country Status (2)

Country Link
JP (1) JPWO2022123968A1 (en)
WO (1) WO2022123968A1 (en)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2013231641A (en) * 2012-04-27 2013-11-14 Toyo Univ Spectroscopic substrate
US20130327129A1 (en) * 2011-02-28 2013-12-12 Moon Chul Jung Spectroscopic sample inspection for automated chromatography
WO2014027652A1 (en) * 2012-08-17 2014-02-20 独立行政法人科学技術振興機構 Method and device for biomolecule analysis using raman spectroscopy

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20130327129A1 (en) * 2011-02-28 2013-12-12 Moon Chul Jung Spectroscopic sample inspection for automated chromatography
JP2013231641A (en) * 2012-04-27 2013-11-14 Toyo Univ Spectroscopic substrate
WO2014027652A1 (en) * 2012-08-17 2014-02-20 独立行政法人科学技術振興機構 Method and device for biomolecule analysis using raman spectroscopy

Also Published As

Publication number Publication date
JPWO2022123968A1 (en) 2022-06-16

Similar Documents

Publication Publication Date Title
US7411673B2 (en) Scanning spectrophotometer for high throughput fluorescence detection
JP6258353B2 (en) Optical measurement apparatus and method for analyzing samples contained in droplets
US8729502B1 (en) Simultaneous, single-detector fluorescence detection of multiple analytes with frequency-specific lock-in detection
JP5920996B2 (en) System and method for detecting and imaging a two-dimensional sample array
US7576855B2 (en) Spectrophotometric method and apparatus
KR101790825B1 (en) Quantum-yield measurement device
US20070177149A1 (en) Instrumentation and method for optical measurement of samples
JPH10170429A (en) Adjustable excitation and/or adjustable detection microplate reader
JP2007501414A (en) Multi-dimensional fluorescence apparatus and method for rapid and highly sensitive quantitative analysis of mixtures
US5422719A (en) Multi-wave-length spectrofluorometer
KR20150037977A (en) Dual spectrometer
JP2011513740A (en) Time-resolved spectroscopic analysis method and system using photon mixing detector
KR20170052256A (en) Apparatus and method for measuring concentration of material
JP2004117298A (en) Measuring method and apparatus using total reflection attenuation
WO2022123968A1 (en) Raman analysis plate, raman analysis device, analysis system, and raman analysis method
JP2000283960A (en) Micro-chip electrophoretic device
US20110122412A1 (en) Devices and methods for optical detection
JP2006153460A (en) Fluorescence detection method, detection device and fluorescence detection program
US7446867B2 (en) Method and apparatus for detection and analysis of biological materials through laser induced fluorescence
JP5208774B2 (en) Fluorescence spectrophotometer
KR100691528B1 (en) Instrument of scanning surface plasmon microscope for the analysis of protein arrays
KR20000051127A (en) Multifunctional Spectrophotometer with CCD detector and Integrating Sphere
JP2004527767A (en) Optical detection of chemical species in enriched media
KR100728897B1 (en) Dual function surface plasmon resonance biosensor
EP4206656A1 (en) Method for enhancing a raman contribution in a spectrum, spectroscopy system and non-transitory computer-readable storage medium

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 21903072

Country of ref document: EP

Kind code of ref document: A1

ENP Entry into the national phase

Ref document number: 2022568109

Country of ref document: JP

Kind code of ref document: A

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 21903072

Country of ref document: EP

Kind code of ref document: A1