Liquid chromatograph and analytical method

Abstract

A liquid chromatograph includes: a sample injection flow path; an analytical pump that guides the mobile phase to the sample injection flow path; a sample injection unit that guides the sample to the sample injection flow path; a flow path switching unit; a separation column that separates the sample that has passed through the sample injection flow path; and a detector that detects the sample that has passed through the separation column, wherein the sample injection flow path includes: a first confluence connected to the sample injection unit and the analytical pump; a metering flow path; and a first branch.

Description

Technical Field

[0001] This invention relates to liquid chromatographs and analytical methods. Background Technology

[0002] Extracolumn diffusion refers to the phenomenon where the sample diffuses within the mobile phase outside the column due to mixing with the mobile phase. When extracolumn diffusion is significant, peak widths increase, preventing the column from achieving its maximum separation capability. To suppress extracolumn diffusion, it is crucial to suppress diffusion in the piping and other components of the instrument. Patent Document 1 describes a liquid chromatograph that suppresses extracolumn diffusion by reducing the internal volume of the flow path around the injection port. Particularly in nano-liquid chromatography (Nano LC), where the flow rate of the mobile phase is relatively low, it is desirable to reduce the cross-sectional area of ​​the flow path, such as piping, proportionally to the flow rate.

[0003] Patent Document 1: Japanese Patent Application Publication No. 2009-276355 Summary of the Invention

[0004] The problem the invention aims to solve

[0005] On the other hand, even if the cross-sectional area of ​​the flow path, such as the piping, is reduced—for example, the groove width (flow path width) of the injection valve rotor can only be reduced to about 50–100 μm due to machining limitations. Therefore, in a liquid chromatograph equipped with an injection valve, the sample flows through a relatively wide flow path formed within the valve. In this case, the contact area between the sample flowing within the flow path and its inner surface increases, or turbulence occurs when flowing from a narrower flow path to a wider one, thus increasing extra-column diffusion. Furthermore, as another problem, when a sample initially at atmospheric pressure is connected to a high-pressure analytical flow path via the injection valve switching action, the pressure in the analytical flow path temporarily drops sharply, causing pressure fluctuations. Further, the injection valve switching action generates wear powder from the rotor, which is typically made of resin, potentially becoming foreign matter and clogging the flow path. If a filter is installed downstream of the injection valve to remove this foreign matter, extra-column diffusion at the filter section increases. Due to these problems, there is a risk of decreased analytical accuracy when analyzing samples passing through valves and similar structures.

[0006] The purpose of this invention is to provide a liquid chromatograph and analytical method that can analyze samples with higher precision.

[0007] Solution for solving the problem

[0008] A first aspect of the present invention relates to a liquid chromatograph, comprising: a sample injection flow path; an analytical pump for guiding a mobile phase to the sample injection flow path; a sample injection unit for guiding a sample to the sample injection flow path; a flow path switching unit; a separation column for separating the sample that has passed through the sample injection flow path; and a detector for detecting the sample that has passed through the separation column. The sample injection flow path includes: a first confluence connected to the sample injection unit and the analytical pump; a metering flow path disposed downstream of the first confluence; and a first branch disposed downstream of the metering flow path and connected to the sample injection unit. The flow path switching unit switches between a first state and a second state, wherein in the first state, a sample is guided from the sample injection unit to the sample injection flow path via the first confluence, and the sample is filled into the metering flow path by delivering the sample flowing through the first branch to the sample injection unit; and in the second state, the sample filled in the metering flow path is delivered to the separation column by guiding the mobile phase from the analytical pump to the first confluence.

[0009] A second aspect of the present invention relates to an analytical method applied in a liquid chromatograph, the liquid chromatograph comprising: a sample injection flow path; an analytical pump for guiding a mobile phase into the sample injection flow path; a sample injection section for guiding a sample into the sample injection flow path; a flow path switching section for switching the flow path; a separation column for separating the sample that has passed through the sample injection flow path; and a detector for detecting the sample that has passed through the separation column. The sample injection flow path includes: a first confluence connected to the sample injection section and the analytical pump; a metering flow path disposed downstream of the first confluence; and a first branch disposed downstream of the metering flow path and connected to the sample injection section. The analytical method includes: guiding a mobile phase from the analytical pump to the first confluence; applying pressure to the sample in the sample injection section; guiding the sample from the sample injection section via the first confluence to the sample injection path, and filling the metering path with the sample by delivering the sample flowing through the first branch to the sample injection section; and discharging the sample filled in the metering path to the separation column by guiding the mobile phase from the analytical pump to the first confluence.

[0010] The effects of the invention

[0011] According to the present invention, samples can be analyzed with higher precision. Attached Figure Description

[0012] [ Figure 1 ] Figure 1 This is a schematic diagram illustrating the configuration of a liquid chromatograph according to an embodiment of the present invention.

[0013] [ Figure 2 ] Figure 2 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0014] [ Figure 3 ] Figure 3 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0015] [ Figure 4 ] Figure 4 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0016] [ Figure 5 ] Figure 5 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0017] [ Figure 6 ] Figure 6 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0018] [ Figure 7 ] Figure 7 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0019] [ Figure 8 ] Figure 8 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0020] [ Figure 9 ] Figure 9 This is a schematic diagram showing a modified example of a liquid chromatograph.

[0021] [ Figure 10 ] Figure 10 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0022] [ Figure 11 ] Figure 11 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0023] [ Figure 12 ] Figure 12 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0024] [ Figure 13 ] Figure 13 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure.

[0025] [ Figure 14 ] Figure 14 This is a schematic diagram of a liquid chromatograph used to illustrate an example of an analytical procedure. Detailed Implementation

[0026] Hereinafter, the analysis system of the present invention will be described in detail with reference to the accompanying drawings.

[0027] (1) Analysis of the system composition

[0028] Figure 1 This is a schematic diagram illustrating the configuration of a liquid chromatograph according to an embodiment of the present invention. Figure 1 The liquid chromatograph 1 is a liquid chromatograph with a low flow rate of mobile phase. For example, the liquid chromatograph 1 is a nano-liquid chromatograph (Nano LC) with a mobile phase flow rate of 1 μL / min. The inner diameter of the tubing constituting the liquid chromatograph 1 is set in the range of 10 to 50 μm. The liquid chromatograph 1 includes a dispensing unit 10, a sample injection unit 20, a flow path switching unit 30, a sample injection flow path 40, a separation column 50, a detector 60, and a control unit 70.

[0029] The infusion unit 10 includes a first analytical pump 11, a second analytical pump 12, and an infusion pressure sensor 13. The first analytical pump 11 is connected to piping P11, and the second analytical pump 12 is connected to piping P12. The first analytical pump 11 delivers liquid stored in container CA to piping P11. The second analytical pump 12 delivers liquid stored in container CB to piping P12. In this embodiment, container CA stores water, and container CB stores an organic solvent. The organic solvent is, for example, methanol, acetonitrile, or a mixture thereof with water. The infusion pressure sensor 13 measures the water pressure within piping P1, which is supplied from the first analytical pump 11. The first and second analytical pumps 11 and 12 may internally include flow meters and flow feedback mechanisms (neither shown), and feedback control can be performed based on flow meter measurements to achieve accurate flow rates.

[0030] The sample injection unit 20 includes a first metering pump 21, a sample pressure sensor 22, a sample aspiration unit 23, a second metering pump 24, and a sample switching valve VA. The sample aspiration unit 23 includes a needle 232, a tube 233, a metering loop 234, and an inlet 235. The needle 232 is connected to one end of the metering loop 234 via the tube 233. The other end of the metering loop 234 is connected to the first metering pump 21. The sample pressure sensor 22 is provided in the flow path connecting the metering loop 234 and the first metering pump 21. The needle 232 can be moved horizontally and vertically via a moving mechanism (not shown). The first metering pump 21 is, for example, a syringe pump. The first metering pump 21 can switch between a state where liquid is drawn from the needle 232 of the sample aspiration unit 23 toward the metering loop 234, and a state where liquid is guided from the tubing P23 to the sample aspiration unit 23. With these configurations, the sample aspiration unit 23 can draw a sample from a container containing the analyte sample into the metering loop 234. Furthermore, the sample aspiration unit 23 can discharge the sample held in the metering loop 234 to the injection port 235. The injection port 235 is connected to one end of the piping P21. The second metering pump 24 is connected to both piping P24 and piping P25. The second metering pump 24 is, for example, a syringe pump, which guides liquid from piping P25 to piping P24.

[0031] The sample switching valve VA is a rotary valve with ports aa1 to ff1. The sample switching valve VA can switch between the VAA state (connecting ports aa1 and bb1, cc1 and dd1, and ee1 and ff1) and the VAB state (connecting ports bb1 and cc1, dd1 and ee1, and ff1 and aa1).

[0032] Port aa1 is connected to container CC via pipe P22. A one-way valve ch1 is provided on pipe P22. Container CC contains the same liquid as either container CA or container CB. In this embodiment, container CC contains the same water as container CA. Although not shown, a switching valve device can be provided on pipe P22 to allow selection of different solvents from multiple containers CC for aspiration. Port bb1 is connected to the first metering pump 21 via pipe P23. Ports cc1 and dd1 are sealed by sealing members SM1 and SM2. Port ee1 is connected to the second metering pump 24 via pipe P24. Port ff1 is connected to one end of pipe P26. The other end of pipe P26 is connected to a waste liquid mechanism (not shown) outside the liquid chromatograph 1. Furthermore, a one-way valve ch2 is provided on pipe P26. Thus, the liquid flowing through pipe P26 is discharged to the waste liquid mechanism outside the liquid chromatograph 1.

[0033] The flow path switching unit 30 includes a first switching valve V1 and a second switching valve V2. The first switching valve V1 is a rotary valve having ports a1 to f1. The first switching valve V1 can switch between a V1A state (connecting ports a1 and b1, c1 and d1, and e1 and f1) and a V1B state (connecting ports b1 and c1, d1 and e1, and f1 and a1). Port b1 is connected to the first analytical pump 11 via piping P11. Port e1 is connected to the second analytical pump 12 via piping P12.

[0034] Port c1 is connected to mixer TA via pipe P31. Pipe P31 is equipped with a porous or mesh filter FT1, which has one side intersecting the liquid flow direction and the other side. Therefore, since foreign matter in the liquid flowing through pipe P31 is collected by filter FT1, the risk of foreign matter clogging the small inner diameter flow path is reduced. Port d1 is connected to mixer TA via pipe P32. Pipe P32 is equipped with filter FT2. Filter FT2 has the same configuration as filter FT1.

[0035] The mixer TA is also connected to piping PA. When the first switching valve V1 is in the V1B state, port b1 is connected to port c1, and port e1 is connected to port d1. Thus, water guided from the first analytical pump 11 to piping P31 and organic solvent guided from the second analytical pump 12 to piping P32 are combined at the mixer TA. The mixer TA is, for example, a gradient mixer that mixes water contained in container CA and organic solvent contained in container CB. Furthermore, the mixture within the mixer TA is guided as the mobile phase to piping PA.

[0036] Port f1 is connected to mixer TB via pipe P33. Filter FT3 is installed on pipe P33. Filter FT3 has the same configuration as filter FT1. Port a1 is connected to mixer TB via pipe P34. Filter FT4 is installed on pipe P34. Filter FT4 has the same configuration as filter FT1.

[0037] The mixer TB is also connected to piping PB. When the first switching valve V1 is in the V1A state, port b1 is connected to port a1, and port e1 is connected to port f1. Thus, water guided from the first analytical pump 11 to piping P31 and organic solvent guided from the second analytical pump 12 to piping P32 are combined at the mixer TB. The mixer TB is, for example, a gradient mixer that mixes water contained in container CA and organic solvent contained in container CB. Furthermore, the mixture within the mixer TB is guided as the mobile phase to piping PB.

[0038] The second switching valve V2 is a rotary valve with ports a2 to f2. The second switching valve V2 can switch between a V2A state (connecting ports a2 and b2, c2 and d2, and e2 and f2) and a V2B state (connecting ports b2 and c2, d2 and e2, and f2 and a2). Port a2 is connected via piping P21 to the inlet 235 of the sample aspiration section 23 of the sample injection section 20. Port b2 is connected via piping PC to the first confluence TC of the sample injection flow path 40 (described later). A filter FT5 is provided on piping PC. The filter FT5 has the same configuration as filter FT1. Port c2 is connected via piping PD to the first branch TD of the sample injection flow path 40 (described later). Port d2 is connected via piping P25 to the second metering pump 24. Port e2 is connected via piping P26 to port f2.

[0039] The sample injection flow path 40 includes a first confluence TC, a metering flow path PP, a first branch TD, and a second confluence TE. The first confluence TC is connected via piping PC to port b2 of the second switching valve V2 and piping PA. The first confluence TC is also connected to one end of the metering flow path PP. The first confluence TC is, for example, a T-shaped branch pipe. The metering flow path PP is a pipe with the same one end and the other end as piping P11, but its length can be adjusted appropriately as needed. For example, the metering flow path PP can be lengthened by winding it between one end and the other end.

[0040] The first branch section TD is connected to port c2 of the second switching valve V2 via pipe PD, and further connected to the second confluence section TE via pipe P41. The first branch section TD is, for example, a T-shaped branch pipe, guiding the mixture of sample and mobile phase flowing through the metering flow path PP to port c2 of the second switching valve V2 and the second confluence section TE. The second confluence section TE is connected to pipe PB and connected to the separation column 50 via pipe P42. The second confluence section TE is, for example, a T-shaped branch pipe. The sample guided from pipe P41 to the second confluence section TE is guided to the detector 60 via the separation column 50. In addition, the mobile phase guided from pipe PB to the second confluence section TE is guided to the detector 60 via the separation column 50. The detector 60 is, for example, a mass spectrometer, which sequentially detects the sample components eluted from the separation column 50.

[0041] The control unit 70 is composed of, for example, a CPU (Central Processing Unit), a ROM (Read-Only Memory), a RAM (Random Access Memory), and a storage device. The ROM or storage device included in the control unit 70 stores an analysis program for performing the analysis operations described later. The control unit 70 is implemented by executing a computer program, such as the analysis program stored in the ROM or storage device, on the RAM. Part or all of the control unit 70 may be constructed from hardware such as electronic circuits. Based on the analysis program, the control unit 70 performs the following analysis operations by controlling the structures of the infusion unit 10, the sample injection unit 20, the flow path switching unit 30, and the detector 60.

[0042] (2) Analyze the action

[0043] Figures 2-8 This is a schematic diagram of a liquid chromatograph 1 used to illustrate an example of analytical operation. Figures 2-8 In the diagram, piping for liquid flow is indicated by thick lines. Analytical procedures include equilibration, sample aspiration, pre-compression, filling, analytical preparation, analytical processing, and cutoff gradient processing. These procedures, as shown below, are executed under the control of the control unit 70.

[0044] In analyzing the motion, the first step is to perform a balancing process. Figure 2 The state of liquid chromatograph 1 during the equilibration process is shown. (Example:) Figure 2 As shown, during the balancing process, the first switching valve V1 of the flow path switching unit 30 switches to state V1A, and the second switching valve V2 of the flow path switching unit 30 switches to state V2B. In this state, the first analytical pump 11 and the second analytical pump 12 operate. Consequently, the water stored in container CA is guided to mixer TB sequentially through pipe P11, ports b1 and a1 of the first switching valve V1, and pipe P34 and filter FT4. Furthermore, the organic solvent stored in container CB is guided to mixer TB sequentially through pipe P12, ports e1 and f1 of the first switching valve V1, and pipe P33 and filter FT3.

[0045] In mixer TB, water and organic solvent are mixed in a predetermined ratio. The mixed liquid is guided to pipe PB as the mobile phase. The mobile phase guided to pipe PB passes sequentially through the second confluence TE of sample injection flow path 40, pipe P42, and separation column 50 before being guided to detector 60. The sum of the flow rates of the first analytical pump 11 and the second analytical pump 12 is the flow rate of the mobile phase supplied to separation column 50. Furthermore, the ratio of the flow rates of the first analytical pump 11 and the second analytical pump 12 is the mixing ratio of the mobile phase supplied to separation column 50. Thus, a mobile phase with a predetermined flow rate and mixing ratio is supplied to separation column 50. The pressure of pipe P11 measured by infusion pressure sensor 13 at this time is referred to as the first pressure below. Thus, in the equilibration process, the mobile phase is supplied to separation column 50 before the analytical processing described later.

[0046] In this embodiment, the sample injection unit 20 is cleaned during the equilibration process. While cleaning the sample injection unit 20, the sample switching valve VA of the sample injection unit 20 is switched to the VAA state. Furthermore, with the needle 232 of the sample aspiration unit 23 connected to the inlet 235, the first metering pump 21 or the second metering pump 24 reciprocates. In this case, water stored in the container CC is sequentially guided to the needle 232 via pipe P22, ports aa1 and bb1 of the sample switching valve VA, pipe P23, metering loop 234, and tube 233. Additionally, water injected from the needle 232 into the inlet 235 is sequentially guided to pipe P26 via pipe P21, ports a2 and f2 of the second switching valve V2, pipe P26, ports e2 and d2 of the second switching valve V2, pipe P25, pipe P24, and ports ee1 and ff1 of the sample switching valve VA. Furthermore, the water flowing through each flow path in the sample injection unit 20 is discharged to a waste liquid mechanism (not shown) outside the liquid chromatograph 1. Therefore, while cleaning each flow path in the sample injection unit 20, clean liquid (water) is filled into each flow path in the sample injection unit 20.

[0047] Next, sample aspiration and processing will be carried out. Figure 3 The status of liquid chromatograph 1 during sample aspiration and processing is shown. (Example) Figure 3 As shown, during sample aspiration, the sample aspiration unit 23 performs an aspiration action. Here, a moving mechanism (not shown) moves the needle 232 along with the tube 233. After moving the needle 232 above the sample container containing the analyte sample, the needle 232 is lowered. This inserts the tip of the needle 232 into the sample container. In this state, the first metering pump 21 operates. As a result, the analyte sample is aspirated from the needle 232 toward the first metering pump 21, and the sample is held within the metering loop 234. After the sample is held, the needle 232 is moved to the injection port 235. In this embodiment, equilibration is performed during the sample aspiration process.

[0048] Next, a pre-compression treatment is performed. Pre-compression treatment refers to the process of applying pressure to the sample in advance. Figure 4 The state of the liquid chromatograph 1 during pre-compression is shown. During pre-compression, the second switching valve V2 of the flow path switching unit 30 is switched to the V2B state, and the sample switching valve VA of the sample injection unit 20 is switched to the VAB state. In addition, with the needle 232 of the sample aspiration unit 23 connected to the injection port 235, either the first metering pump 21 or the second metering pump 24 operates.

[0049] In this configuration, in the flow path of the sample injection section 20, with the downstream of port dd1 sealed by the sealing member SM2, the first metering pump 21 or the second metering pump 24 operates, thereby applying pressure to the sample within the flow path of the sample injection section 20. Feedback control is performed on the first metering pump 21 or the second metering pump 24 based on the pressure of the water and the sample (hereinafter referred to as the sample) flowing through each pipe within the sample injection section 20, as measured by the sample pressure sensor 22. This adjusts the pressure of the sample within the sample injection section 20. In this embodiment, the pressure of the sample within the sample injection section 20 is set to a first pressure. That is, during the pre-pressurization process, the pressure of the sample within the sample injection section 20 is adjusted to the pressure of the mobile phase passing through the separation column 50 during the equilibration process.

[0050] Next, we will perform the filling process. Figure 5 The status of liquid chromatograph 1 during the packing process is shown. (Example:) Figure 5 As shown, the aforementioned balancing process is performed simultaneously during the filling process. During the filling process, the second switching valve V2 of the flow path switching unit 30 is switched to state V2A. Since the pressure of the sample in the sample injection unit 20 has been pre-pressurized to the first pressure, diffusion of the sample in the sample injection unit 20 and backflow to the sample injection unit 20 caused by pressure fluctuations during the switching of the switching valve V2 can be suppressed. Next, the metering pump 21 performs a discharge operation at a predetermined flow rate, while the metering pump 24 performs a suction operation at the same flow rate as the metering pump 21, thereby guiding the sample in the sample injection unit 20, which has been adjusted to the first pressure during the pre-pressurization process, to the sample injection flow path 40 via the piping PC. The sample guided to the sample injection flow path 40 is sequentially guided to the first confluence TC, the metering flow path PP, and the first branch TD, and then guided to the piping P25 via the ports c2 and d2 of the second switching valve V2 via the piping PD. Thus, the sample is guided into the metering flow path PP while maintaining the pressure at the first pressure.

[0051] Figure 6The system is in a state awaiting analysis. During this waiting state, the second switching valve V2 of the flow path switching unit 30 switches to the V2B state, and the sample switching valve VA of the sample injection unit 20 switches to the VAA state. At this time, metering pump 21 or metering pump 24 operates, supplying water from container CC to the sample injection unit 20, and simultaneously guiding the sample from the sample injection unit 20 to piping P26. Consequently, the water flowing through each piping in the sample injection unit 20 is discharged to a waste liquid unit (not shown) outside the liquid chromatograph 1. In other words, the sample injection unit 20 is cleaned during the analysis preparation process. This, for example, removes sample residue from the needle 232.

[0052] The sample, under initial pressure, is contained within the metering flow path PP and subjected to analytical processing. Simultaneously with the start of analytical processing, the detector 60 begins analyzing the sample. Figure 7 The status of liquid chromatograph 1 during analytical processing is shown. For example... Figure 7 As shown, the sample injection section 20 described above can also be cleaned simultaneously during the analytical processing. During the analytical processing, the first switching valve V1 of the flow path switching section 30 is switched to the V1B state. In this case, the water stored in container CA is sequentially guided to mixer TA via pipe P11, ports b1 and c1 of the first switching valve V1, pipe P31, and filter FT1. Furthermore, the organic solvent stored in container CB is sequentially guided to mixer TA via pipe P12, ports e1 and d1 of the first switching valve V1, pipe P32, and filter FT2. In mixer TA, water and organic solvent are mixed in a predetermined ratio. The mixed liquid is then guided to pipe PA as the mobile phase. The mobile phase guided to pipe PA is then guided to metering flow path PP via the first confluence section TC of sample injection flow path 40. Thus, the sample in metering flow path PP is ejected by the mobile phase. The sample and mobile phase within the metering flow path PP are guided to the separation column 50 via the first branch TD, pipe P41, the second confluence TE, and pipe P42. The sample passing through the separation column 50 is then guided to the detector 60.

[0053] Once the sample in the metering flow path PP is guided to the separation column 50, a cutoff gradient treatment is performed. Figure 8 The status of liquid chromatograph 1 during the cutoff gradient process is shown. (Example:) Figure 8As shown, the sample injection section 20 described above can also be cleaned simultaneously during the cutoff gradient processing. During the cutoff gradient processing, the first switching valve V1 of the flow path switching section 30 is switched to state V1A. In this case, the mobile phase guided to the piping PB passes sequentially through the second confluence section TE of the sample injection flow path 40, the piping P42, and the separation column 50 before being guided to the detector 60. In this state, the composition (ratio of water to organic solvent) of the mobile phase guided to the separation column 50 is changed by controlling the first analytical pump 11 and the second analytical pump 12, thereby performing gradient analysis. Furthermore, the mobile phase is guided to the separation column 50 without passing through the metering flow path PP. In this case, since the flow path from the analytical pumps 11 and 12 to the separation column 50 can be shortened (cutoff) during gradient analysis, the mobile phase with a changed composition can reach the separation column 50 within a shorter flow path, thereby shortening the analysis time.

[0054] According to the liquid chromatograph 1 of the above embodiment, the mobile phase is guided to the sample injection path 40 through the infusion unit 10, and the sample is guided to the sample injection path 40 through the sample injection unit 20. When the second switching valve V2 of the flow path switching unit 30 is in the V2A state, after the sample is guided from the sample injection unit 20 to the first confluence TC of the sample injection path 40, a portion of the sample returns from the first branch TD of the sample injection path 40 to the sample injection unit 20. In this case, the sample is filled in the sample injection unit 20, the first confluence TC, the metering path PP, and the first branch TD. In this state, when the first switching valve V1 of the flow path switching unit 30 changes to the V1B state, the mobile phase is guided from the infusion unit 10 to the first confluence TC. Thus, since the mobile phase is guided from the first confluence TC toward the separation column 50, the sample in the metering path PP located between the first confluence TC and the separation column 50 can be guided to the separation column 50. Thus, according to the configuration of the present invention, when the sample located in the metering flow path PP is delivered and analyzed through the infusion section 10, the sample is directly guided to the separation column without passing through the valve groove, which is difficult to reduce in diameter, thereby suppressing extra-column diffusion of the sample. As a result, the sample can be analyzed with higher precision. Furthermore, the metering flow path PP is initially connected to the infusion section 10 from the confluence section TE side, thereby maintaining the same pressure as during delivery. In addition, the sample is also pre-pressurized to a state equal to the pressure of the metering flow path PP. Therefore, when the state of the switching valve V2 is switched, the sample injection section 20 is connected to the metering flow path PP, and the pre-pressurized sample is introduced into the metering flow path PP, pressure fluctuations and the resulting diffusion are suppressed. In this state, the sample in the metering flow path PP is delivered to the separation column.

[0055] Furthermore, when the first switching valve V1 of the flow path switching section 30 is in state V1A, the mobile phase is guided from the infusion section 10 to the second confluence section TE of the sample injection flow path 40. In this case, since the second confluence section TE is located closer to the separation column 50 than the first confluence section TC, the metering flow path PP, and the first branch section TD, the mobile phase is guided to the separation column from a position closer to the separation column 50. As a result, in gradient analysis, the flow path from the analytical pumps 11 and 12 to the separation column 50 can be shortened (cut off), allowing the mobile phase with a changed composition to reach the separation column 50 within a shorter flow path, thereby shortening the analysis time. Moreover, the sample portion filled in the piping PC and PD that is not guided to the separation column 50 can be prevented from diffusing from the confluence section TC or the branch section TD and gradually flowing into the separation column 50, thus preventing unnecessary baseline changes.

[0056] Furthermore, first filters FT1 and FT2 are provided between the first switching valve V1 of the flow path switching section 30 and the first junction TC of the sample injection flow path 40, and third filters FT3 and FT4 are provided between the first switching valve V1 and the second junction TE. In this case, foreign matter such as dust generated due to wear in the first switching valve V1 can be removed. Similarly, since filter FT5 is provided between the second switching valve V2 of the flow path switching section 30 and the first junction TC of the sample injection flow path 40, foreign matter such as dust generated due to wear in the second switching valve V2 can be removed. Thus, especially at nanoliter flow rates, the risk of flow paths with small inner diameters being blocked by foreign matter can be reduced.

[0057] (3) Variations

[0058] Figure 9 This is a schematic diagram illustrating a modified liquid chromatograph 1A. The liquid chromatograph 1 of the above embodiment differs from the liquid chromatograph 1A in the following aspects: The liquid chromatograph 1A includes a sample injection unit 20A instead of a sample injection unit 20. The sample injection unit 20A includes a first sample pump 25, a second sample pump 26, a mixer TF, and a sample switching valve VB.

[0059] A first sample pump 25 is connected to a mixer TF via piping P27, and a second sample pump 26 is connected to the mixer TF via piping P28. The first sample pump 25 delivers liquid stored in container CD to piping P27, and the second sample pump 26 delivers liquid stored in container CE to piping P28. In this embodiment, container CD contains the same water as container CA, and container CE contains the same organic solvent as container CB. The mixer TF mixes the water stored in container CD with the organic solvent stored in container CE and directs the mixture to piping P29. The first sample pump 25 or the second sample pump 26 is equipped with a second pressure sensor (not shown) capable of measuring the pressure of the delivered liquid.

[0060] The sample switching valve VB is a rotary valve with ports aa2 to ff2. The sample switching valve VB can switch between the VBA state (connecting ports aa2 and bb2, cc2 and dd2, and ee2 and ff2) and the VBB state (connecting ports bb2 and cc2, dd2 and ee2, and ff2 and aa2).

[0061] Port aa2 is connected to the inlet 235 of the sample aspiration unit 23, port bb2 is connected to the waste liquid flow path, and port cc2 is connected to the aspiration unit 21A via pipe P30. The aspiration unit 21A is, for example, a syringe. In addition, port dd2 is connected to the metering loop 234 via pipe P31, port ee2 is connected to the mixer TF via pipe P29, and port ff2 is connected to port a2 of the second switching valve V2 of the flow path switching unit 30 via pipe P21.

[0062] Furthermore, ports e2 and f2 of the second switching valve V2 of the flow path switching unit 30 are connected via pipe PE1. Port d2 of the second switching valve V2 of the flow path switching unit 30 is connected to the resistance tube RT via pipe PE2. The resistance tube RT has a smaller diameter than the other pipes constituting the liquid chromatograph 1A. This allows for an increase in the pressure of the liquid in the flow path connected to the resistance tube RT. To ensure that the pressure of the liquid in the flow path is the same as the pressure of the mobile phase supplied to the separation column 50 during the equilibration process, the inner diameter of the resistance tube RT can be arbitrarily set. For example, the inner diameter of the resistance tube RT can be appropriately selected in the range of 50 to 100 μm. In this embodiment, the inner diameter of the resistance tube RT is set to an inner diameter that allows the pressure of the liquid in the flow path connected to the resistance tube RT to reach a first pressure. The other end of the resistance tube RT is connected to a waste liquid mechanism (not shown) outside the liquid chromatograph 1A.

[0063] (4) Analysis actions in the variant examples

[0064] Figures 10-14 This is a schematic diagram of a liquid chromatograph 1A used to illustrate an example of analytical operation. Figures 10-14 In the diagram, piping for liquid flow is indicated by thick lines. The analysis in the variant example performs the same processing as the analysis described above.

[0065] Figure 10 The state of liquid chromatograph 1A during the equilibration process is shown. The equilibration process is the same as that for liquid chromatograph 1 described above. Therefore, as... Figure 10As shown, in the equilibration process, water stored in container CA is directed to mixer TB, and organic solvent stored in container CB is directed to mixer TB. In mixer TB, the liquid resulting from the mixture of water and organic solvent is directed to pipe PB as the mobile phase. The mobile phase directed to pipe PB is then directed to detector 60 after passing sequentially through the second confluence TE of sample injection path 40, pipe P42, and separation column 50.

[0066] During the equilibration process, the sample injection unit 20A is cleaned. While cleaning the sample injection unit 20A, the sample switching valve VB of the sample injection unit 20 is switched to the VBB state, and the second switching valve V2 is switched to the V2B state. Furthermore, with the needle 232 of the sample aspiration unit 23 connected to the injection port 235, the first sample pump 25 and the second sample pump 26 operate. In this situation, water stored in container CD and organic solvent stored in container CE are guided to the sample aspiration unit 23 via ports ee2, dd2 of the sample switching valve VB and pipe P31. Water guided to the sample aspiration unit 23 is then guided to pipe P21 via ports aa2, ff2. Additionally, water in pipe P21 is sequentially guided to pipe PE2 via ports a2, f2 of the second switching valve V2, pipe PE1, and ports e2, d2 of the second switching valve V2. Water guided to pipe PE2 is discharged through the resistance tube RT to a waste liquid mechanism (not shown) outside the liquid chromatograph 1. Thus, while cleaning each flow path within the sample injection section 20, clean liquid (water) is filled into each flow path within the sample injection section 20. Furthermore, by connecting each pipe of the sample injection section 20 to the resistance tube RT, the first sample pump 25 and the second sample pump 26 are indirectly connected to the resistance tube RT, thereby changing the liquid in each pipe within the sample injection section 20 to the first pressure.

[0067] Next, sample aspiration and processing will be carried out. Figure 11 The state of the liquid chromatograph 1A during sample aspiration is shown. During sample aspiration, the sample switching valve VB of the sample injection unit 20 is switched to the VBA state. The aspiration operation of the sample aspiration unit 23 during sample aspiration is the same as described above. Furthermore, with the needle 232 inserted into a sample container (not shown), the aspiration unit 21A operates. Thus, the analyte sample is aspirated from the needle 232 toward the aspiration unit 21A, thereby holding the sample within the quantitative loop 234. Equilibration continues during sample aspiration.

[0068] Next, pre-compression treatment is performed. Figure 12 The status of liquid chromatograph 1A during pre-compression treatment is shown. (Example:) Figure 12As shown, the aforementioned balancing process is performed simultaneously during the pre-compression treatment. During the pre-compression treatment, the sample switching valve VB of the sample injection unit 20A is switched to the VBB state. Furthermore, with the needle 232 of the sample aspiration unit 23 connected to the inlet 235, the first sample pump 25 and the second sample pump 26 are activated for liquid delivery. In this case, the sample held in the metering loop 234 is guided to the piping P21. Since the first sample pump 25 and the second sample pump 26 are indirectly connected to the resistance tube RT, feedback control is performed on the flow rate of the first sample pump 25 or the second sample pump 26, ensuring that the pressure of the sample flowing through the sample injection unit 20, measured by the second pressure sensor mounted on the pump, equals the first pressure.

[0069] Next, we will perform the filling process. Figure 13 The status of liquid chromatograph 1A during the packing process is shown. (Example) Figure 13 As shown, the aforementioned balancing process is performed simultaneously during the filling process. During the filling process, the second switching valve V2 of the flow path switching unit 30 is switched to the V2A state. In this case, the sample in the piping P21, which has undergone pressure adjustment during the pre-compression process, is guided to the sample injection flow path 40 via the piping PC. The sample guided to the sample injection flow path 40 is sequentially guided to the first confluence TC, the metering flow path PP, and the first branch TD, then guided to the piping PE2 via the piping PD, and subsequently to the resistance pipe RT. In this case, the sample, which has been adjusted to the first pressure during the pre-compression process, is guided to the metering flow path PP of the sample injection flow path 40. Furthermore, since the metering flow path PP of the sample injection flow path 40 is indirectly connected to the resistance pipe RT, the pressure within the metering flow path PP is maintained. Thus, while the pressure within the metering flow path PP is maintained at the first pressure, the sample is guided to the metering flow path PP. Therefore, fluctuations in the pressure applied to the sample are suppressed.

[0070] Subsequently, analysis and preparation processing were carried out. Figure 14 The status of the liquid chromatograph 1A during analytical preparation is shown. (Example:) Figure 14 As shown, during the analytical preparation process, the aforementioned balancing process and the cleaning of the sample injection section 20A are performed simultaneously. During the analytical preparation process, the second switching valve V2 of the flow path switching section 30 is switched to the V2B state. In this case, the sample supply to the sample injection flow path 40 stops, and therefore the sample at the first pressure is contained within the metering flow path PP.

[0071] The sample, under the initial pressure, is contained within the metering flow path PP and subjected to analytical processing. Simultaneously with the start of analytical processing, the sample is analyzed via detector 60. Regarding analytical processing, due to... Figure 7The operation is the same, so it will be omitted here. After the sample in the metering flow path PP is guided to the separation column 50, a cutoff gradient treatment is performed. Regarding the cutoff gradient treatment, since it is similar to... Figure 8 The same applies, so the explanation is omitted here.

[0072] According to the liquid chromatograph 1A of the above embodiment, the sample injection unit 20 is connected to the resistance tube RT. In this case, the pressure inside the sample injection unit 20 can be increased. Therefore, the sample pressure can be pre-increased inside the sample injection unit 20 before the sample is guided to the metering flow path PP. As a result, when the sample is guided to the metering flow path PP, fluctuations in sample pressure are suppressed, thereby suppressing extra-column diffusion.

[0073] (5) Other implementation methods

[0074] In the above embodiment, filters FT1 and FT2 are provided between the first switching valve V1 and the mixer TA in the flow path switching section 30, and filters FT3 and FT4 are provided between the first switching valve V1 and the mixer TB. However, the present invention is not limited thereto. For example, filters FT1 and FT2 can be replaced by a new filter between the mixer TA and the first confluence section TC. Similarly, filters FT3 and FT4 can be replaced by a new filter between the mixer TB and the second confluence section TE.

[0075] (6) The correspondence between the constituent elements of the claims and the parts of the embodiments

[0076] The following describes examples of the correspondence between the constituent elements of the claims and the elements of the embodiments. Filters FT1 and FT2 are examples of first filters, filters FT3 and FT4 are examples of second filters, and filter FT5 is an example of a third filter. Furthermore, mixer TA is an example of a first mixing section, mixer TB is an example of a second mixing section, and the first sample pump 25 and the second sample pump 26 are examples of filling pumps.

[0077] (7) Method

[0078] Those skilled in the art will understand that the above-described exemplary embodiments are specific examples of the following approaches.

[0079] (Item 1) A liquid chromatograph relating to a method includes:

[0080] Sample injection flow path;

[0081] The analytical pump directs the mobile phase into the sample injection path;

[0082] The sample injection section guides the sample into the sample injection flow path;

[0083] Flow path switching unit;

[0084] A separation column for separating the sample that has passed through the sample injection flow path; and

[0085] The detector measures the sample that has passed through the separation column.

[0086] The sample injection path includes:

[0087] The first confluence section is connected to the sample injection section and the analytical pump;

[0088] The first branch is connected to the separation column and the flow path switching section; and

[0089] A metering flow path is located between the first confluence and the first branch.

[0090] The flow path switching unit switches between a first state and a second state, wherein,

[0091] In the first state, the sample is filled from the sample injection section into the metering flow path via the first confluence section;

[0092] The second state involves guiding the mobile phase from the analytical pump to the first confluence, thereby delivering the sample filled in the metering flow path to the separation column.

[0093] According to the liquid chromatograph described in claim 1, the mobile phase is guided to the sample injection path by an analytical pump, and the sample is guided to the sample injection path through the sample injection unit. When the flow path switching unit is in the first state, after the sample is guided from the sample injection unit to the sample injection path, a portion of the sample returns from the first branch of the sample injection path to the sample injection unit. In this case, since the sample fills the sample injection unit, the first confluence, the metering path, and the first branch, at least a portion of the sample can be positioned within the metering path. In this state, when the flow path switching unit changes to the second state, the mobile phase is guided from the analytical pump to the first confluence. Thus, since the mobile phase is guided from the first confluence toward the separation column, the sample located in the metering path between the first confluence and the separation column can be guided to the separation column. Therefore, when the sample in the metering path is pumped and analyzed by the analytical pump, it is guided directly to the separation column without passing through a valve groove that is difficult to narrow, thereby suppressing extra-column diffusion of the sample. As a result, sample analysis can be performed with higher precision.

[0094] Furthermore, since the above-mentioned effects are achieved through switching of the flow path switching unit, it is not necessary to stop the operation of the analytical pump from the start to the end of the analysis. Therefore, since the pressure within the flow path constituting the liquid chromatograph does not decrease, there is no need to separately set up an action to increase the pressure within the flow path of the liquid chromatograph from the start to the end of the analysis. Thus, the detection throughput of the liquid chromatograph can be increased.

[0095] (Item 2) In the liquid chromatograph described in Item 1,

[0096] The sample injection flow path further includes: a second confluence portion, disposed between the first branch portion and the separation column.

[0097] The flow path switching unit is able to guide the mobile phase from the analytical pump to the second confluence unit in the first state.

[0098] According to the liquid chromatograph described in item 2, when the flow path switching unit is in the first state, the mobile phase is guided from the analytical pump to the second confluence of the sample injection flow path. In this case, since the second confluence is located closer to the separation column than the first confluence, the metering flow path, and the first branch, the mobile phase is guided from the second confluence to the separation column. Therefore, when the flow path switching unit is in the first state (filled state), the pressure of the mobile phase flowing through the separation column can be adjusted. Thus, since the pressure of the mobile phase flowing through the separation column is pre-adjusted, fluctuations in the mobile phase pressure within the separation column during analysis are suppressed. As a result, extra-column diffusion can be suppressed.

[0099] (Item 3) The liquid chromatograph described in item 2 may further include:

[0100] A first pressure sensor measures the pressure of the flowing phase directed from the analytical pump; and

[0101] The second pressure sensor measures the pressure of the sample guided from the sample injection section to the sample injection flow path.

[0102] According to the liquid chromatograph described in item 3, the difference between the pressure of the mobile phase flowing through the second confluence and the pressure of the sample filling the sample injection section, the first confluence, the metering flow path, and the first branch section can be reduced based on the values ​​measured by the first and second pressure sensors. In this case, when the flow path switching section switches from the first state to the second state, the pressure fluctuation applied to the sample when the sample and mobile phase merge in the first and second confluences is suppressed. Therefore, extracolumn diffusion of the sample can be further suppressed using a simpler configuration.

[0103] (Item 4) In the liquid chromatograph described in Item 2,

[0104] The flow path switching unit may further include:

[0105] A first switching valve, connecting the analytical pump to both the first and second junctions, is capable of switching between a state where the analytical pump is connected to the first junction and a state where the analytical pump is connected to the second junction; and

[0106] The second switching valve can switch between a state in which the sample injection section is connected to the first confluence and the first branch, and a state in which the sample injection section is not connected to the first confluence and the first branch.

[0107] According to the liquid chromatograph described in item 4, the first switching valve can be set to connect the analytical pump and the second confluence, and the second switching valve can be set to connect the sample injection section and the first confluence and the first branch section. In this case, the mobile phase can be guided from the second confluence to the separation column while the sample is filled into the sample injection section, the first confluence, the metering flow path, and the first branch section. At this time, at least a portion of the sample can be positioned within the metering flow path. Alternatively, the first switching valve can be set to connect the analytical pump and the first confluence, and the second switching valve can be set to not connect the sample injection section and the first confluence and the first branch section. This allows at least a portion of the sample within the metering flow path to be guided to the separation column. Thus, with a simple configuration, at least a portion of the sample located in the metering flow path can be guided to the separation column without passing through valves or similar components, thereby suppressing extra-column diffusion of the sample guided to the separation column.

[0108] (Item 5) The liquid chromatograph described in Item 4 may further include:

[0109] A first filter is disposed between the first switching valve and the first junction; and

[0110] The second filter is located between the first switching valve and the second junction.

[0111] According to the liquid chromatograph described in item 5, foreign matter such as dust generated due to wear in the first switching valve can be removed before being guided to the separation column.

[0112] (Item 6) The liquid chromatograph described in Item 5 may further include:

[0113] The third filter is located between the second switching valve and the first junction.

[0114] According to the liquid chromatograph described in item 6, foreign matter such as dust generated due to wear in the second switching valve can be removed before being guided to the separation column.

[0115] (Item 7) In the liquid chromatograph described in Item 2,

[0116] The mobile phase consists of a mixture of a first liquid and a second liquid.

[0117] The analytical pump includes:

[0118] A first analytical pump delivers the first liquid; and

[0119] The second analytical pump delivers the second liquid.

[0120] The liquid chromatograph may further include:

[0121] A first mixing section, disposed between the first analytical pump and the second analytical pump and the first confluence section, mixes the first liquid guided from the first analytical pump and the second liquid guided from the second analytical pump; and

[0122] The second mixing section is located between the first analytical pump, the second analytical pump, and the second confluence section, and mixes the first liquid guided from the first analytical pump and the second liquid guided from the second analytical pump.

[0123] According to the liquid chromatograph described in item 7, after guiding the sample in the metering flow path to the separation column with the mobile phase guided from the first mixing section to the first confluence section, the mobile phase can be guided from the second mixing section to the second confluence section. In this case, the mobile phase guided to the second confluence section is not passed through the first confluence section of the injection flow path, the metering flow path, and the first branch section, but is directly guided to the separation column from the second confluence section, thus enabling cutoff during gradient elution.

[0124] (Item 8) In the liquid chromatographs described in items 1 through 7,

[0125] The sample injection unit includes:

[0126] Quantitative loop;

[0127] The needle is connected to the metering loop; and

[0128] A first metering pump, connected to the side of the metering loop opposite to the needle, is capable of drawing the sample from the needle into the metering loop and discharging the sample from the metering loop back into the needle.

[0129] When the flow path switching unit is in the second state, the sample is drawn from the needle to the metering loop by the first metering pump.

[0130] When the flow path switching section is in the first state, the sample held in the quantitative loop can be guided to the sample injection flow path via the first confluence section.

[0131] According to the liquid chromatograph described in item 8, the sample is drawn into the quantitative loop while the mobile phase is guided to the separation column via the first confluence. Thus, the sample is drawn into the quantitative loop while the analytical pump adjusts the pressure of the mobile phase flowing through the separation column, thereby shortening the series of actions of the liquid chromatograph compared to situations where these actions are performed separately. This, in turn, increases the detection throughput of the liquid chromatograph.

[0132] (Item 9) In the liquid chromatograph described in Item 8,

[0133] The sample injection unit may further include:

[0134] The injection port allows the needle to be inserted and connected to the flow path switching unit;

[0135] A second metering pump is connected to the flow path switching unit; and

[0136] A sealing member that seals the opposite side of the needle of the first metering pump and the opposite side of the flow path switching section of the second metering pump.

[0137] When the flow path switching unit is in the second state, after the sample is drawn from the needle to the quantitative loop by the first metering pump, with the needle inserted into the injection port, the first metering pump performs a discharge action to perform a pre-compression action so that the pressure of the sample in the quantitative loop is equal to the pressure of the mobile phase in the separation column. When the flow path switching unit is in the first state, the first metering pump discharges the sample at a predetermined flow rate, and at the same time, the second metering pump draws the sample at the predetermined flow rate to fill the sample injection flow path.

[0138] According to the liquid chromatograph described in claim 9, the sample is aspirated into the metering loop while the mobile phase is guided to the separation column via the first confluence. Furthermore, after sample aspiration, pressure can be applied to the sample using a first metering pump. Therefore, pressure can be pre-applied to the sample injection section before the sample is guided into the metering flow path.

[0139] (Item 10) The liquid chromatograph described in Item 4 may further include:

[0140] A resistance tube increases the pressure within the connected flow path.

[0141] The resistance pipe is connected to the second switching valve.

[0142] The state in which the second switching valve is not connected to the sample injection section and the first confluence section and the first branch section may include the state in which the sample injection section is connected to the resistance tube.

[0143] According to the liquid chromatograph described in item 10, when the second switching valve is not connected to the sample injection section and the first confluence and first branch sections, the sample injection section is connected to the resistance tube. In this case, the pressure inside the sample injection section can be increased. Therefore, the sample can be pre-pressurized in the sample injection section before being guided into the metering flow path. As a result, fluctuations in sample pressure are suppressed when the sample is guided to the separation column, thereby suppressing extra-column diffusion.

[0144] (Item 11) In the liquid chromatograph described in Item 11,

[0145] The sample injection unit may include:

[0146] Quantitative loop;

[0147] The needle is connected to the metering loop;

[0148] The aspiration section draws the sample from the needle toward the metering loop;

[0149] The injection port allows the needle to be inserted.

[0150] A fill pump directs liquid to the metering loop; and

[0151] The sample switching valve can switch between a state in which the metering loop is connected to the aspiration section and a state in which the filling pump is connected to the metering loop and the injection port is connected to the second switching valve.

[0152] According to the liquid chromatograph described in item 11, the sample can be drawn into the quantitative loop while the sample switching valve is connected to the quantitative loop and the aspirator. Furthermore, after the needle is inserted into the injection port, the sample switching valve can be positioned to connect the packing pump and the quantitative loop, and the injection port and the second switching valve. In this case, the sample in the quantitative loop is guided to the second switching valve by liquid drawn from the packing pump. Moreover, when the second switching valve is positioned to connect the sample injection port and the resistance tube, pressure can be pre-applied to the sample before it is guided into the metering flow path. Furthermore, the above series of actions are performed while the mobile phase is guided to the separation column via the first confluence. Therefore, the detection throughput of the liquid chromatograph is increased.

[0153] (Item 12) Other analytical methods involved, applied in a liquid chromatograph, said liquid chromatograph comprising:

[0154] Sample injection flow path;

[0155] An analytical pump that directs the mobile phase into the sample injection path;

[0156] Guide the sample to the sample injection section of the sample injection flow path;

[0157] Flow path switching unit for switching flow paths;

[0158] A separation column for separating samples that have passed through the sample injection flow path; and

[0159] A detector for detecting samples that have passed through the separation column.

[0160] The sample injection path includes:

[0161] The first confluence section is connected to the sample injection section and the analytical pump;

[0162] The first branch connected to the separation column and the flow path switching section; and

[0163] A metering flow path is provided between the first confluence and the first branch.

[0164] The analytical method includes:

[0165] The step of guiding the mobile phase from the analytical pump to the first confluence;

[0166] The step of applying pressure to the sample within the sample injection section;

[0167] The step of filling the sample from the sample injection section into the metering flow path via the first confluence section; and

[0168] The step of delivering the sample filled in the metering flow path to the separation column by guiding the mobile phase from the analytical pump to the first confluence.

[0169] According to the analytical method described in item 1, pressure can be applied to the sample in the sample injection section while the pressure of the mobile phase flowing through the separation column is adjusted by the analytical pump. Then, the pressure-adjusted sample in the sample injection section is filled into the metering flow path. In this state, the pressure-adjusted sample in the metering flow path is delivered to the separation column by the pressure-adjusted mobile phase. Thus, pressure fluctuations applied to the sample are suppressed when it is guided to the separation column. As a result, extra-column diffusion can be suppressed, thereby enabling more precise sample analysis.

Claims

1. A liquid chromatograph, comprising: Sample injection flow path; The analytical pump directs the mobile phase into the sample injection path; The sample injection section guides the sample into the sample injection flow path; Flow path switching unit; A separation column is used to separate the sample that has passed through the sample injection flow path; as well as The detector measures the sample that has passed through the separation column. The sample injection path includes: The first confluence section is connected to the sample injection section and the analytical pump; The first branch is connected to the separation column and the flow path switching section; as well as A metering flow path is located between the first confluence and the first branch. The flow path switching unit switches between a first state and a second state, wherein, In the first state, the sample is filled from the sample injection section into the metering flow path via the first confluence section; The second state involves guiding the mobile phase from the analytical pump to the first confluence, thereby delivering the sample filled in the metering flow path to the separation column.

2. The liquid chromatograph according to claim 1, characterized in that, The sample injection flow path further includes: a second confluence portion, disposed between the first branch portion and the separation column. In the first state, the flow path switching unit guides the mobile phase from the analytical pump to the second confluence unit.

3. The liquid chromatograph according to claim 2, characterized in that, Further features include: A first pressure sensor measures the pressure of the flowing phase directed from the analytical pump; and The second pressure sensor measures the pressure of the sample guided from the sample injection section to the sample injection flow path.

4. The liquid chromatograph according to claim 2, characterized in that, The flow path switching unit further includes: A first switching valve connects the analytical pump to the first junction and the second junction, and is capable of switching between a state where the analytical pump is connected to the first junction and a state where the analytical pump is connected to the second junction; as well as The second switching valve can switch between a state in which the sample injection section is connected to the first confluence and the first branch, and a state in which the sample injection section is not connected to the first confluence and the first branch.

5. The liquid chromatograph according to claim 4, characterized in that, Further features include: A first filter is disposed between the first switching valve and the first junction; and The second filter is located between the first switching valve and the second junction.

6. The liquid chromatograph according to claim 5, characterized in that, Further features include: The third filter is located between the second switching valve and the first junction.

7. The liquid chromatograph according to claim 2, characterized in that, The mobile phase consists of a mixture of a first liquid and a second liquid. The analytical pump includes: A first analytical pump delivers the first liquid; and The second analytical pump delivers the second liquid. The liquid chromatograph further comprises: A first mixing section is provided between the first analytical pump and the second analytical pump and the first confluence section, for mixing the first liquid guided from the first analytical pump and the second liquid guided from the second analytical pump; as well as The second mixing section is located between the first analytical pump, the second analytical pump, and the second confluence section, and mixes the first liquid guided from the first analytical pump and the second liquid guided from the second analytical pump.

8. The liquid chromatograph according to any one of claims 1 to 7, characterized in that, The sample injection unit includes: Quantitative loop; The needle is connected to the metering loop; and A first metering pump, connected to the side opposite the needle of the metering loop, is capable of drawing the sample from the needle into the metering loop and discharging the sample from the metering loop back into the needle. When the flow path switching unit is in the second state, the sample is drawn from the needle to the metering loop by the first metering pump. When the flow path switching section is in the first state, the sample held in the quantitative loop is guided to the sample injection flow path via the first confluence section.

9. The liquid chromatograph according to claim 8, characterized in that, The sample injection unit further includes: The injection port allows the needle to be inserted and connected to the flow path switching unit; A second metering pump is connected to the flow path switching unit; and A sealing member that seals the opposite side of the needle of the first metering pump and the opposite side of the flow path switching section of the second metering pump. When the flow path switching unit is in the second state, after the sample is drawn from the needle to the quantitative loop by the first metering pump, with the needle inserted into the injection port, the first metering pump performs a discharge action to perform a pre-compression action so that the pressure of the sample in the quantitative loop is equal to the pressure of the mobile phase in the separation column. When the flow path switching unit is in the first state, the first metering pump discharges the sample at a predetermined flow rate, and at the same time, the second metering pump draws the sample at the predetermined flow rate to fill the sample injection flow path.

10. The liquid chromatograph according to claim 4, characterized in that, Further features include: A resistance tube increases the pressure within the connected flow path. The resistance pipe is connected to the second switching valve. The state in which the second switching valve is not connected to the sample injection section and the first confluence section and the first branch section includes the state in which the sample injection section is connected to the resistance tube.

11. The liquid chromatograph according to claim 10, characterized in that, The sample injection unit includes: Quantitative loop; The needle is connected to the metering loop; The aspiration section draws the sample from the needle toward the metering loop; The injection port allows the needle to be inserted; A fill pump directs the liquid to the metering loop; as well as The sample switching valve can switch between a state in which the metering loop is connected to the aspiration section and a state in which the filling pump is connected to the metering loop and the injection port is connected to the second switching valve.

12. An analytical method applied in a liquid chromatograph, the liquid chromatograph comprising: Sample injection flow path; An analytical pump that directs the mobile phase into the sample injection path; Guide the sample to the sample injection section of the sample injection flow path; Flow path switching unit for switching flow paths; A separation column for separating samples that have passed through the sample injection flow path; as well as A detector for detecting samples that have passed through the separation column. The sample injection path includes: The first confluence section is connected to the sample injection section and the analytical pump; A first branch connected to the separation column and the flow path switching section; as well as A metering flow path is provided between the first confluence and the first branch. The analytical method includes: The step of guiding the mobile phase from the analytical pump to the first confluence; The step of applying pressure to the sample within the sample injection section; The step of filling the sample from the sample injection section into the metering flow path via the first confluence section; and The step of delivering the sample filled in the metering flow path to the separation column by guiding the mobile phase from the analytical pump to the first confluence.

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