Concentration device, analysis system including same, and concentration method

Abstract

To provide a concentration device that can concentrate an object component in liquid in a short time and can recover a small amount of liquid containing the post-concentrated object component, to provide an analysis system that includes the concentration device, and to provide a concentration method.SOLUTION: A concentration device 10 comprises: a housing 20; a separation membrane ME that partitions an internal space of the housing 20 so as to form a flow pass FP in the housing 20; a first supply part that supplies a first liquid from a first position of the housing 20 to the flow pass FP so that the first liquid flows in a first direction along the separation membrane ME; a second supply part that supplies a second liquid from a second position of the housing 20 to the flow pass FP so that the second liquid flows in a second direction opposite the first direction along the separation membrane ME; and a third supply part that supplies a third liquid containing an object component having a size not transmitted through the separation membrane ME from a third position of the housing 20 to the flow pass FP, in which the third position is present between the first position and second position in the first direction.SELECTED DRAWING: Figure 4

Description

[Technical Field]

[0001] The present invention relates to a concentrating device, an analytical system including the same, and a concentrating method. [Background technology]

[0002] Tangential flow filtration (TFF) is sometimes used for purifying viruses and concentrating or desalting proteins, peptides, nucleic acids, etc. For example, the virus production method described in Patent Document 1 involves concentrating a stabilized virus harvest by tangential flow filtration.

[0003] In a concentration device using tangential flow filtration, a solution containing the component to be concentrated (hereinafter referred to as the target component) is flowed along a separation membrane. The pore size of the separation membrane is smaller than that of the target component. Pore ​​diameter Components smaller than 100 μm permeate the separation membrane, while the target component does not. The solution containing the target component is collected in a reservoir and then made to flow again along the separation membrane. In this way, the solution containing the target component circulates through a flow path including the separation membrane and the reservoir, thereby concentrating the target component in the solution. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Patent No. 5548207 Summary of the Invention [Problem to be solved by the invention]

[0005] However, in conventional concentrating devices using tangential flow filtration, the above-described solution circulation must be repeated to concentrate the target component in the solution, which consumes a considerable amount of time. Furthermore, the total volume of the solution containing the target component after concentration is relatively large. This makes handling a solution containing a certain amount of the target component complicated. Therefore, it is desirable to recover a small amount of solution containing a certain amount of the target component.

[0006] An object of the present invention is to provide a concentrating device that can concentrate a target component in a liquid in a short period of time and recover a small amount of liquid containing the target component after concentration, an analytical system equipped with the same, and a concentration method. [Means for solving the problem]

[0007] A concentration device according to one aspect of the present invention comprises a housing, a separation membrane that partitions the internal space of the housing to form a flow path within the housing, a first supply unit that supplies a first liquid from a first position in the housing to the flow path so that the first liquid flows in a first direction along the separation membrane, a second supply unit that supplies a second liquid from a second position in the housing to the flow path so that the second liquid flows in a second direction opposite to the first direction along the separation membrane, and a third supply unit that supplies a third liquid containing a target component having a size that does not permeate the separation membrane to the flow path from a third position in the housing, the third position being between the first position and the second position in the first direction.

[0008] A concentration method according to another aspect of the present invention includes the steps of: supplying a first liquid from a first position in a housing to a flow path along a separation membrane in the housing so that the first liquid flows in a first direction along the separation membrane; supplying a second liquid from a second position in the housing to the flow path so that the second liquid flows in a second direction opposite to the first direction along the separation membrane in the housing; and supplying a third liquid containing a target component having a size that does not permeate the separation membrane to the flow path from a third position in the housing, the third position being between the first position and the second position in the first direction. [Effects of the Invention]

[0009] According to the present invention, it is possible to concentrate a target component in a liquid in a short time, and to recover a small amount of liquid containing the target component after concentration. [Brief explanation of the drawings]

[0010] [Figure 1] 1 is a schematic diagram illustrating a configuration of an analysis system including a concentration device according to an embodiment. [Figure 2] FIG. 10 is a diagram showing an example of a chromatogram generated based on an output signal from a detector of a liquid chromatograph. [Figure 3] 2 is a timing chart showing the transition of states of each part in the analysis system of FIG. 1. [Figure 4] 2 is a schematic diagram showing an accumulation operation in the concentration operation of the analysis system of FIG. 1. FIG. [Figure 5] FIG. 2 is a schematic diagram showing a recovery operation in a concentration operation of the analysis system of FIG. [Figure 6] FIG. 1 shows chromatograms obtained in Example 1 and Comparative Example 1. [Figure 7] FIG. 1 shows a chromatogram obtained in Example 2. DETAILED DESCRIPTION OF THE INVENTION

[0011] Hereinafter, a concentration device, an analysis system, and a concentration method according to embodiments will be described in detail with reference to the drawings.

[0012] (1) Analysis system configuration 1 is a schematic diagram showing the configuration of an analytical system including a concentrating device according to one embodiment. The analytical system 100 includes a liquid chromatograph 1 and a concentrating device 10. The liquid chromatograph 1 includes a pump 3, an autosampler 4, a separation column 5, a detector 6, and pipes p11 to p15.

[0013] In this embodiment, the discharge port of the pump 3 is connected to an autosampler 4 through a pipe p11. The autosampler 4 is connected to an inlet of a separation column 5 through a pipe p12. The outlet of the separation column 5 is connected to an inlet of a detector 6 through a pipe p13.

[0014] A switching valve V3 is provided at the outlet of the detector 6. The switching valve V3 has one liquid inlet, a first liquid outlet, and a second liquid outlet. The outlet of the detector 6 is connected to the liquid inlet of the switching valve V3 through a pipe p14. A pipe p15 is connected to the first liquid outlet of the switching valve V3. The state in which the liquid inlet of the switching valve V3 is connected to the first liquid outlet is called the first state. The state in which the liquid inlet of the switching valve V3 is connected to the second liquid outlet is called the second state.

[0015] An eluent serving as a mobile phase is contained in the eluent container 2. The pump 3 draws the eluent from the eluent container 2 and guides the drawn eluent to a separation column 5 through a pipe p11, an autosampler 4, and a pipe p12.

[0016] The autosampler 4 introduces the sample to be analyzed into the eluent guided from the pump 3 to the separation column 5. This introduces the eluent containing the sample into the separation column 5. The separation column 5 separates the sample in the eluent into its components over time. The eluent containing the separated sample is introduced from the outlet of the separation column 5 through a pipe p13 to the inlet of the detector 6. The detector 6 detects the components of the separated sample. The detector 6 is, for example, a UV (ultraviolet) detector. A chromatogram is generated based on the output signal of the detector 6. Hereinafter, these operations will be referred to as the analytical operation. The eluent containing the sample guided from the detector 6 is then guided to the switching valve V3 through a pipe p14. When the switching valve V3 is switched to the first state, the eluent guided to the switching valve V3 is guided to a waste tank or the like (not shown) through a pipe p15.

[0017] In this embodiment, a concentrating device 10 is used to concentrate one or more components in an eluent delivered from a detector 6 of a liquid chromatograph 1. The concentrating device 10 includes a housing 20, a control unit 30, pumps 8a and 8b, a fraction collector F1, a flow rate adjustment valve V1, switching valves V3 and V4, an on-off valve V5, and pipes p1 to p5. The housing 20 is formed in a cylindrical shape having an internal space extending in one direction Y, with one end and the other end being closed. The cross-sectional shape of the housing 20 may be circular, elliptical, polygonal, or any other shape.

[0018] A separation membrane ME is provided in the internal space of the housing 20 in parallel with one direction Y so as to partition the space into a flow path FP and a discharge space SP. In this embodiment, an ultrafiltration membrane is used as the separation membrane ME. The ultrafiltration membrane has a pore size of about 0.001 μm to 0.01 μm. Here, a separation membrane having a pore size smaller than the size of the target component to be concentrated is used as the separation membrane ME. In this embodiment, a porous sintered body 21 is provided in the discharge space SP to support the separation membrane ME. Note that a mesh-structured sieve may be provided instead of the porous sintered body 21.

[0019] First to third supply ports P1 to P3 and first and second discharge ports P4, P5 are formed in the housing 20 of the concentrating apparatus 10. The first to third supply ports P1 to P3 and the first discharge port P4 are formed to communicate between a flow path FP within the housing 20 and the outside of the housing 20.

[0020] In this embodiment, the first supply port P1 is provided at one end of the flow path FP of the housing 20, and the first discharge port P4 is provided at the other end of the flow path FP of the housing 20. In this embodiment, the first supply port P1 and the first discharge port P4 are on the same straight line.

[0021] Hereinafter, the direction from the first supply port P1 to the first discharge port P4 will be referred to as the first direction a, and the direction from the first discharge port P4 to the first supply port P1 will be referred to as the second direction b. The first direction a and the second direction b are opposed to each other. In this embodiment, the first direction a and the second direction b are parallel to one direction Y. Note that the first supply port P1 and the first discharge port P4 do not have to be on the same straight line, and the first direction a and the second direction b do not have to be on a straight line. In other words, the first direction a and the second direction b may form another angle close to 180°.

[0022] The second supply port P2 is provided at a position between the first supply port P1 and the first discharge port P4 in the direction Y. Furthermore, the third supply port P3 is provided at a position between the first supply port P1 and the second supply port P2 in the direction Y. In this embodiment, the third supply port P3 is provided at a position closer to the first supply port P1 than the second supply port P2 in the direction Y. The second discharge port P5 is formed to communicate between the discharge space SP in the housing 20 and the outside of the housing 20.

[0023] The first supply port P1 of the housing 20 is connected to the outlet of the pump 8a through a pipe p1 and a flow rate control valve V1. The pump 8a draws the eluent contained in the eluent container 7a and guides it to the first supply port P1 through the flow rate control valve V1 and the pipe p1. As a result, the eluent is supplied from the first supply port P1 into the flow path FP of the housing 20, and the eluent supplied from the first supply port P1 flows in a first direction a along the separation membrane ME. The flow rate control valve V1 is configured to be able to switch the flow rate of the eluent delivered to the pipe p1 between a first flow rate and a second flow rate greater than the first flow rate. Hereinafter, the state in which the flow rate control valve V1 delivers the eluent to the pipe p1 at the first flow rate will be referred to as the first state, and the state in which the eluent is delivered at the second flow rate will be referred to as the second state.

[0024] The second supply port P2 of the housing 20 is connected to the discharge port of the pump 8b through a pipe p2. The pump 8b sucks the eluent contained in the eluent container 7b and guides it to the second supply port P2 through the pipe p2. As a result, the eluent is supplied from the second supply port P2 into the flow path FP of the housing 20, and flows along the separation membrane ME in the second direction b and also in the first direction a. In this embodiment, the eluent contained in the eluent containers 7a and 7b is the same as the eluent contained in the eluent container 2.

[0025] The third supply port P3 of the housing 20 is connected to the second liquid outlet of the switching valve V3 through a pipe p3. When the switching valve V3 is switched to the second state, the eluent delivered from the detector 6 is guided through the pipe p3 to the third supply port P3 of the housing 20. As a result, the eluent containing the target component is supplied from the third supply port P3 toward the separation membrane ME of the flow path FP of the housing 20.

[0026] The switching valve V4 has a liquid inlet, a first liquid outlet, and a second liquid outlet. A first discharge port P4 of the housing 20 is connected to the liquid inlet of the switching valve V4 through a pipe p4. A first liquid outlet of the switching valve V4 is connected to a fraction collector (FRC) F1 through a pipe p16. The eluent in the flow path FP of the housing 20 is discharged from the first discharge port P4 through the pipe p4 to the liquid inlet of the switching valve V4. When the switching valve V4 is switched to the first state, the eluent in the flow path FP of the housing 20 is guided to the fraction collector F1 through the pipe p16. When the switching valve V4 is switched to the second state, the eluent in the flow path FP of the housing 20 is guided to a waste tank or the like (not shown) through a pipe p17.

[0027] One or more sample containers SC are arranged in the fraction collector F1. The fraction collector F1 collects a certain amount of eluent containing a target component, which is guided from the flow path FP of the housing 20, into the sample container SC.

[0028] Among the multiple components in the eluent flowing along the separation membrane ME in the flow path FP of the housing 20, components having a size smaller than the pore size of the separation membrane ME permeate the separation membrane ME together with the eluent. The eluent and components that permeate the separation membrane ME from the flow path FP are guided to the discharge space SP of the housing 20. The second discharge port P5 of the housing 20 is connected to the on-off valve V5 through a pipe p5. When the on-off valve V5 is opened, the eluent in the discharge space SP is discharged through the pipe p5 to a waste liquid tank or the like (not shown).

[0029] The control unit 30 controls the operations of the pump 3, autosampler 4, detector 6, flow rate adjustment valve V1 of the concentrating device 10, switching valves V3 and V4, on-off valve V5, pumps 8a and 8b, and fraction collector F1. Thus, in this embodiment, the control unit 30 controls the components of the liquid chromatograph 1 and the concentrating device 10. Note that a control unit that controls the liquid chromatograph 1 and a control unit that controls the concentrating device 10 may be provided separately.

[0030] The control unit 30 includes a CPU (Central Processing Unit), a ROM (Read Only Memory), a RAM (Random Access Memory), etc. The ROM stores in advance an analysis program for executing the analysis operation of the liquid chromatograph 1, a concentration program for executing the concentration operation of the concentrator 10 (described later), and the elution time of the target component. The analysis operation and concentration operation are performed by the CPU executing the analysis program and concentration program stored in the ROM on the RAM.

[0031] Fig. 2 shows an example of a chromatogram generated based on the output signal from the detector 6 of the liquid chromatograph 1. The horizontal axis of the chromatogram represents elution time (retention time), and the vertical axis represents signal intensity. In the chromatogram shown in Fig. 2, multiple peaks A to C appear, which represent multiple components in the sample that have been separated over time by the separation column 5.

[0032] (2) Operation of the Analysis System 100 The analysis system 100 according to this embodiment performs an analysis operation using a liquid chromatograph 1 and a concentration operation using a concentration device 10. The concentration operation includes an accumulation operation and a recovery operation. The accumulation operation is an operation in which the target component in the eluent is collected at a predetermined location on the separation membrane ME. The recovery operation is an operation in which the target component collected by the accumulation operation is discharged from the housing 20 together with the eluent and collected in a sample container SC.

[0033] Fig. 3 is a timing chart showing the transition of the states of each part in the liquid chromatograph 1 of Fig. 1. Fig. 4 is a schematic diagram showing the accumulation operation in the concentration operation of the analysis system 100. Fig. 5 is a schematic diagram showing the recovery operation in the concentration operation of the analysis system 100. In Figs. 4 and 5, the flow of liquid is indicated by thick lines. Here, a case will be described in which the component corresponding to peak C in the chromatogram of Fig. 2 is concentrated as the target component.

[0034] Figure 3 shows the changes over time in the states of the flow rate adjustment valve V1, the switching valves V3 and V4, the on-off valve V5, and the pumps 8a and 8b during the analysis and concentration operations. In the example of Figure 3, the analysis operation begins at time t1, and the concentration operation is performed from time t2 to time t5. Of the concentration operations, the accumulation operation is performed from time t3 to time t4, and the recovery operation is performed from time t4 to time t5.

[0035] First, the control unit 30 in FIG. 4 switches the switching valve V3 to the first state. As a result, the eluent discharged from the pump 3 is guided to the detector 6 through the pipe p12, autosampler 4, pipe p13, and separation column 5. At this time, the autosampler 4 introduces a sample into the eluent. As a result, the separation column 5 separates the components of the sample in the eluent. In this state, the separated components are detected by the detector 6. The control unit 30 generates, for example, the chromatogram shown in FIG. 2 based on the output signal of the detector 6. The eluent delivered from the detector 6 is discharged to a waste tank or the like through the switching valve V3 and pipe p15. At time t1, the pumps 8a and 8b are off, the switching valve V4 is in the second state, and the on-off valve V5 is closed.

[0036] Next, at a predetermined time t2 from time t1, the control unit 30 turns on the pump 8a and switches the flow rate control valve V1 to the first state. As a result, the eluent in the eluent container 7a is supplied to the flow path FP in the housing 20 through the pump 8a, the pipe p1, and the first supply port P1 of the housing 20. The eluent supplied to the flow path FP through the first supply port P1 flows in the first direction a along the separation membrane ME. The control unit 30 also turns on the pump 8b. As a result, the eluent in the eluent container 7b is supplied to the flow path FP in the housing 20 through the pump 8b, the pipe p2, and the second supply port P2 of the housing 20. The eluent supplied to the flow path FP from the second supply port P2 flows in the first direction a and the second direction b along the separation membrane ME. In this case, the eluent flowing from the first supply port P1 in the first direction a and the eluent flowing from the second supply port P2 in the second direction b collide between the first supply port P1 and the second supply port P2. The control unit 30 also opens the on-off valve V5 to allow the eluent to be discharged from the discharge space SP in the housing 20.

[0037] The accumulation operation starts at time t3 when the detector 6 detects a component corresponding to peak C in FIG. 2 (hereinafter referred to as target component c). At time t3, the control unit 30 switches the switching valve V3 to the second state. As a result, the eluent containing the target component c output from the detector 6 is supplied to the flow path FP in the housing 20 through the switching valve V3, the pipe p3, and the third supply port P3 of the housing 20.

[0038] An eluent containing a target component c is supplied to the collision position between the eluent supplied from the first supply port P1 and flowing in a first direction a and the eluent supplied from the second supply port P2 and flowing in a second direction b.

[0039] 4, the third supply port P3 is disposed at a position closer to the first supply port P1 than the second supply port P2. In this case, the flow rate of the eluent in the pipe p2 is set to be larger than the flow rate of the eluent in the pipe p1 so that the eluent from the third supply port P3 is supplied to a position where the eluent supplied from the first supply port P1 and the eluent supplied from the second supply port P2 collide.

[0040] In this way, an eluent containing target component c is supplied from the third supply port P3 to the collision position between the eluent flowing in the first direction a and the eluent flowing in the second direction b, so that the target component c accumulates on the separation membrane ME. Also, while the target component c is accumulating, a portion of the eluent flowing in the first direction a along the separation membrane ME and a portion of the eluent flowing in the second direction b permeate the separation membrane ME by tangential flow filtration. Furthermore, the eluent supplied from the third supply port P3 is absorbed by the separation membrane ME. Pore ​​diameter Components having a size smaller than 1 permeate the separation membrane ME. The eluent and components that permeate the separation membrane ME are led from the second discharge port P5 through a pipe p5 and an on-off valve V5 to a waste tank or the like (not shown).

[0041] When a predetermined time has elapsed since time t3 when the accumulation operation started, or when the signal strength of the detector 6 becomes equal to or less than a predetermined value, it is determined that accumulation of the target component c has ended, and a recovery operation is started at time t4. At time t4, the control unit 30 switches the switching valve V3 to the first state. This stops the supply of eluent from the third supply port P3 into the flow path FP of the casing 20. The control unit 30 also turns off the pump 8b. This stops the supply of eluent from the second supply port P2 into the flow path FP of the casing 20. The control unit 30 also switches the switching valve V4 to the first state. The control unit 30 also closes the opening / closing valve V5.

[0042] In this state, the control unit 30 switches the flow rate control valve V1 to the second state. This allows the eluent to continue being supplied to the first supply port P1. In this case, the amount of eluent supplied to the first supply port P1 during the collection operation is greater than the amount of eluent supplied to the first supply port P1 during the accumulation operation. This creates a flow of eluent in the first direction a from the first supply port P1 to the first discharge port P4 within the flow path FP of the housing 20. As a result, the eluent in the flow path FP of the housing 20 and the target component c accumulated on the separation membrane ME are guided from the first discharge port P4 through the pipe p4, the switching valve V4, and the pipe p16 to the fraction collector F1. The eluent containing the target component c is then collected in the sample container SC located in the fraction collector F1. The concentration operation by the concentrator 10 ends at time t5 when the eluent containing the target component c is collected in the sample container SC.

[0043] (3) Example 1 and Comparative Example 1 In Example 1, the recovery rate of the target component during the accumulation operation of the concentrator 10 was evaluated. In this Example 1, the flow rates of the first supply port P1, the second supply port P2, the third supply port P3, the first discharge port P4, and the second discharge port P5 of the concentrator 10 were controlled using a pump, syringe, switching valve, mass flow meter, etc. An ultrafiltration membrane with a molecular weight cutoff of 10 kDa was used as the separation membrane ME. A UV (ultraviolet) detector using 280 nm ultraviolet light was connected to the first discharge port P4. BSA (bovine serum albumin) manufactured by Sigma-Aldrich was used as the target component. 1.0 mg of BSA was dissolved in 10 mL of ultrapure water to prepare a BSA solution with a concentration of 0.1 mg / mL.

[0044] During the accumulation operation, a 0.1 mg / mL BSA solution was injected into the flow channel FP from the third supply port P3 using a 1 mL manual injector. Ultrapure water was supplied into the flow channel FP from the first supply port P1 and the second supply port P2. The flow rate of the first supply port P1 was set to 0.5 mL / min, and the flow rate of the second supply port P2 was set to 1.7 mL. The flow rate of the first discharge port P4 was set to 1.2 mL / min, and the flow rate of the second discharge port P5 was set to 1.2 mL / min. This allowed the BSA solution to accumulate on the separation membrane ME for 2.5 minutes.

[0045] During the recovery operation, the flow rate of the first supply port P1 was set to 1.2 mL / min, and the flow rates of the second supply port P2 and the third supply port P3 were set to 0 mL / min. Furthermore, the BSA accumulated on the separation membrane ME was discharged from the first discharge port P4 for approximately 1 minute. At this time, the flow rate of the second discharge port P5 was set to 0 mL / min, and the flow rate of the tangential flow filtration was set to 0 mL / min. The BSA solution discharged from the first discharge port P4 was introduced into a UV detector.

[0046] In Comparative Example 1, no accumulation operation was performed, and a BSA solution with a concentration of 0.1 mg / mL was injected into the flow path FP from the third supply port P3 using a 1 mL manual injector, and the BSA solution discharged from the first discharge port P4 by a recovery operation was led to the UV detector.

[0047] FIG. 6 shows chromatograms obtained by UV detection of the BSA solution recovered after the accumulation operation in Example 1 and the BSA solution recovered without the accumulation operation in Comparative Example 1. In the chromatograms in FIG. 6, peak D of BSA detected in Example 1 is indicated by a solid line, and peak E of BSA detected in Comparative Example 1 is indicated by a dashed line. The area value of peak D was 2,761,087 [μV·sec]. The area value of peak E was 2,808,307 [μV·sec]. The recovery rate of BSA when the accumulation operation was performed was 98.5%, which is almost the same as when the accumulation operation was not performed. This confirms that the recovery rate of the target component is high when the accumulation operation in the concentrator 10 according to this embodiment is used.

[0048] (4) Example 2 In Example 2, two types of solutions containing the same amount of the target component but having different concentrations were subjected to accumulation using the same concentrator 10 as in Example 1, and the signal intensities of the peaks in the chromatograms were compared.

[0049] In Example 2, a BSA solution with a concentration of 1.0 mg / mL and a total volume of 100 μL (hereinafter referred to as the first BSA solution) and a BSA solution with a concentration of 0.1 mg / mL and a total volume of 1 mL (hereinafter referred to as the second BSA solution) were used. Both the first BSA solution and the second BSA solution contained 0.1 mg of BSA.

[0050] The first BSA solution was injected into the flow path FP from the third supply port P3, and an accumulation operation and a collection operation were performed. The time for the accumulation operation of the first BSA solution was set to 1 minute. The second BSA solution was injected into the flow path FP from the third supply port P3, and an accumulation operation and a collection operation were performed. The time for the accumulation operation of the second BSA solution was set to 2.5 minutes. Other conditions for the accumulation operation and the collection operation were the same as those in Example 1.

[0051] FIG. 7 shows a chromatogram obtained in Example 2. In the chromatogram in FIG. 7, peak F, which corresponds to BSA when the first BSA solution was used, is shown by a solid line, and peak G, which corresponds to BSA when the second BSA solution was used, is shown by a dashed line. Note that the elution times of peak F and peak G differ due to different accumulation operation times. In the chromatogram in FIG. 7, the height (signal intensity) of peak F was 104,146 [μV], and the height (signal intensity) of peak G was 98,920 [μV]. In this case, the height of peak G relative to the height of peak F was 95.0%. This result confirmed that when the amount of the target component contained in the solution concentrated by the concentrator 10 is the same, peaks of approximately the same height can be detected even if the concentration of the solution is different.

[0052] (5) Effects of the embodiment According to the concentrating device 10 of this embodiment, the target component c in the eluent is accumulated at a position on the separation membrane ME where the eluent flowing in the first direction a and the eluent flowing in the second direction b collide. While the target component c is being accumulated, a portion of the eluent supplied from the first supply port P1 and the second supply port P2 flowing along the separation membrane ME permeates the separation membrane ME by tangential flow filtration. Furthermore, Eluent supplied from the third supply port P3 Among them, separation membrane ME Pore ​​diameter Components smaller than the size of the target component c permeate the separation membrane ME. In this case, the concentration of the target component c can be increased without circulating the eluent multiple times. Therefore, it is possible to concentrate the target component c in the liquid in a short time and recover a small amount of eluent containing the concentrated target component c. For example, in the concentration operation, the target component c can be concentrated in about 1 to 2 minutes, and in the recovery operation, it is possible to recover 2 mL or less of the eluent containing the target component c.

[0053] In addition, in this embodiment, the position where the eluent collides is set directly below the third supply port P3, so that the target component c supplied from the third supply port P3 is accumulated without being dispersed in the first direction a or the second direction b, thereby improving the accumulation efficiency of the target component c.

[0054] Furthermore, in this embodiment, during the recovery operation, the eluent remaining on the separation membrane ME in the flow path FP is discharged from the first discharge port P4 together with the accumulated target component c. At this time, a portion of the eluent remaining on the separation membrane ME permeates the separation membrane ME by tangential flow filtration. Therefore, the amount of target component accumulated on the separation membrane ME can be increased while the amount of eluent discharged from the first discharge port P4 can be reduced. Furthermore, since the first discharge port P4 is provided at the other end of the housing 20, the amount of eluent permeating the separation membrane ME by tangential flow filtration increases. Therefore, the concentration efficiency of the target component c is improved.

[0055] Furthermore, in this embodiment, the eluent and target component c remaining in the flow path FP during the recovery operation are discharged from the flow path FP by the eluent flowing in the first direction a. Therefore, it is possible to recover the eluent containing the concentrated target component c with simple control.

[0056] Furthermore, according to the analysis system 100 of this embodiment, a specific component in the eluent analyzed by the liquid chromatograph 1 is concentrated by the concentrator 10. This makes it possible to concentrate the target component in the sample analyzed by the liquid chromatograph 1 in a short period of time, and to easily recover a small amount of eluent containing the concentrated target component.

[0057] Furthermore, an eluent containing a target component in the sample is supplied to the third supply port P3. This allows the desired component in the sample analyzed by the liquid chromatograph 1 to be concentrated as a target component c. This makes it possible to concentrate one or more desired components from among the components of the sample analyzed by the liquid chromatograph 1.

[0058] (6) Other embodiments Although the concentrator 10 is included in the analysis system 100, the concentrator 10 may be used alone. In this case, the supply of liquid to the first supply port P1, the second supply port P2, the first discharge port P4, and the second discharge port P5 of the concentrator 10 may be controlled using, for example, a pump, a syringe, a switching valve, a mass flow meter, or the like. The concentrator 10 according to the above embodiment can also be used for concentrating or desalting proteins, peptides, nucleic acids, and the like, as well as for purifying viruses.

[0059] The liquid supplied to the first and second supply ports P1 and P2 is the eluent used in the liquid chromatograph 1, but other liquids, such as separately prepared ultrapure water, may also be used. In this case, it is possible to desalt the eluent and sample supplied from the third supply port P3. Furthermore, connecting a mass spectrometer or the like to the first discharge port P4 allows for structural analysis of the target component c.

[0060] (7) Correspondence between each component of the claims and each part of the embodiment The following describes an example of the correspondence between each component of the claims and each element of the embodiments. In the above embodiment, the flow rate adjustment valve V1, the pump 8a, and the pipe p1 are an example of a first supply unit, the pump 8b and the pipe p2 are an example of a second supply unit, the switching valve V3 and the pipe p3 are an example of a third supply unit, and the switching valve V4 and the pipe p4 are an example of a first discharge unit.

[0061] (8) Mode It will be appreciated by those skilled in the art that the exemplary embodiments described above are examples of the following aspects.

[0062] (Item 1) A concentrating device according to one aspect comprises: The housing and a separation membrane that partitions the internal space of the housing so as to form a flow path within the housing; a first supply unit that supplies a first liquid from a first position in the housing to the flow path so that the first liquid flows in a first direction along the separation membrane; a second supply unit that supplies the second liquid from a second position in the housing to the flow path so that the second liquid flows along the separation membrane in a second direction opposite to the first direction; a third supply unit that supplies a third liquid containing a target component having a size that does not permeate the separation membrane from a third position of the housing to the flow path; The third position is between the first position and the second position in the first direction.

[0063] In one embodiment of the concentrating device, a first liquid flowing in a first direction and a second liquid flowing in a second direction collide along a separation membrane in a flow path between a first position and a second position in the first direction. At this time, a portion of the first liquid and a portion of the second liquid permeate the separation membrane by tangential flow filtration. In this state, a target component in a third liquid supplied to the flow path from a third position between the first and second positions is accumulated at the position where the first liquid and the second liquid collide. While the target component is being accumulated, a portion of the first liquid and a portion of the second liquid flowing along the separation membrane permeate the separation membrane by tangential flow filtration. In addition, a portion of the third liquid permeates the separation membrane by tangential flow filtration. Pore ​​diameter Components having a size smaller than the separation membrane will permeate the separation membrane.

[0064] In this way, the target component accumulates on the separation membrane, and the separation membrane Pore ​​diameter Components of the first, second, and third liquids that are smaller than the particle size of the first, second, and third liquids permeate the separation membrane. Therefore, the concentration of the target component can be increased without circulating the third liquid multiple times. As a result, the target component in the liquid can be concentrated in a short time, and a small amount of liquid containing the concentrated target component can be recovered.

[0065] (Item 2) In the concentrating device described in item 1, the third position may correspond to a position where a flow of the first liquid in the first direction and a flow of the second liquid in the second direction collide.

[0066] According to the concentrating device described in paragraph 2, the target component is supplied to a position where the first liquid and the second liquid collide. As a result, the target component is accumulated without being dispersed in the first or second direction of the first or second liquid. Therefore, the accumulation efficiency of the target component is improved.

[0067] (Item 3) The concentrating device described in item 1 or 2 may further include a first discharge unit that discharges the first, second, and third liquids from a fourth position in the flow path after the second supply unit stops supplying the second liquid and after the third supply unit stops supplying the third liquid.

[0068] According to the concentrating device described in paragraph 3, the first, second, and third liquids remaining on the separation membrane are discharged by the first discharge section together with the accumulated target component. At this time, some of the first, second, and third liquids remaining on the separation membrane permeate the separation membrane by tangential flow filtration. Therefore, the amount of the target component accumulated on the separation membrane can be increased, and the amount of the first, second, and third liquids discharged from the first discharge section can be reduced.

[0069] (Item 4) In the concentrating device according to item 3, The fourth location may be downstream from the second location in the first direction.

[0070] According to the concentrating device described in paragraph 4, the first, second, and third liquids are discharged from a fourth position located downstream of the second position. In this case, the first, second, and third liquids flow along the separation membrane in the first direction and are discharged, so that the amount of the first, second, and third liquids that permeates the separation membrane by tangential flow filtration increases. This improves the concentration efficiency of the target component.

[0071] (Section 5) Article 3 term or fourth The concentrating device according to the item a control unit that controls the first supply unit, the second supply unit, the third supply unit, and the first discharge unit; The control unit controlling the first supply unit, the second supply unit, and the third supply unit so that, during a first operation, the first liquid flows in a first direction in the flow path, the second liquid flows in a second direction, and a third liquid is supplied to the flow path; During the second operation after the first operation, the first, second and third supply units and the first discharge unit may be controlled so that the first liquid is supplied to the flow path so as to flow in the first direction while the second and third liquids are not supplied to the flow path, and the first liquid is discharged from the flow path together with the second and third liquids remaining on the separation membrane.

[0072] According to the concentrating device described in paragraph 5, during the first operation, the target component is accumulated at the position where the first liquid and the second liquid collide. Also, during the second operation, the first, second, and third liquids and the target component remaining in the flow path are discharged from the flow path. In this case, it is possible to recover the liquid containing the concentrated target component with simple control.

[0073] (Item 6) In the concentrating device according to item 5, During a first operation, the first supply unit supplies a first liquid to the flow path at a first flow rate and the second supply unit supplies a second liquid to the flow path at a second flow rate greater than the first flow rate; The third position may be located closer to the first position in the first direction than the second position.

[0074] According to the concentrating device described in paragraph 6, during the first operation, the first liquid flows in a first direction at a first flow rate, and the second liquid flows in a second direction at a second flow rate greater than the first flow rate. As a result, the first liquid and the second liquid collide in the first direction near the first position. As a result, the target component accumulates at a position closer to the first position than the second position. As a result, during the second operation, the first liquid flows from the first position in the first direction and is discharged from the fourth position, thereby increasing the distance along the separation membrane that the first, second, and third liquids remaining on the separation membrane travel. This improves the efficiency of tangential flow filtration. As a result, the concentration efficiency of the target component improves.

[0075] (Section 7) The analysis system is A liquid chromatograph, The apparatus may further include a concentrating device according to any one of items 1 to 6, which concentrates a specific component in the eluent discharged after analysis by the liquid chromatograph as a target component in the third liquid.

[0076] According to the analytical system described in paragraph 7, a specific component in the eluent analyzed by the liquid chromatograph is concentrated in the concentrator, thereby making it possible to concentrate the target component in the liquid in a short time and recover a small amount of liquid containing the concentrated target component.

[0077] (Item 8) In the analysis system according to item 7, The liquid chromatograph is a separation column; a mobile phase supply unit that supplies a mobile phase to the separation column; a sample introduction unit that introduces a sample into a mobile phase to be supplied to a separation column by a mobile phase supply unit; a detector for detecting components of the sample in the mobile phase derived from the separation column; The third supply unit may supply the mobile phase containing the sample discharged from the detector to the flow channel as the third liquid.

[0078] According to the analytical system described in paragraph 8, the mobile phase containing the sample is used as the third liquid, which allows the components of the sample analyzed by the liquid chromatograph to be concentrated as the target component.

[0079] (Section 9) Article 8 In the analysis system according to the item The third supply unit may supply one or more components of the sample discharged from the detector as target components to the flow channel together with the third liquid.

[0080] According to the analytical system described in paragraph 9, it becomes possible to concentrate one or more components of a sample to be analyzed by liquid chromatography.

[0081] (Item 10) A step of supplying a first liquid to a flow path from a first position in the housing so that the first liquid flows in a first direction along the separation membrane in the flow path along the separation membrane in the housing; supplying a second liquid to the flow path from a second position in the housing so that the second liquid flows along the separation membrane in the housing in a second direction opposite to the first direction; supplying a third liquid containing a target component having a size that does not permeate the separation membrane from a third position in the housing to the flow path; The method of concentrating, wherein the third position is between the first position and the second position in the first direction.

[0082] In the concentration method according to the tenth aspect, the target component accumulates on the separation membrane, and the separation membrane Pore ​​diameter Components of the first, second, and third liquids that are smaller than the particle size of the first, second, and third liquids permeate the separation membrane. Therefore, the concentration of the target component can be increased without circulating the third liquid multiple times. As a result, the target component in the liquid can be concentrated in a short time, and a small amount of liquid containing the concentrated target component can be recovered. [Explanation of symbols]

[0083] 1...liquid chromatograph, 2, 7...eluent container, 3, 8...pump, 4...autosampler, 5...separation column, 6...detector, 10...concentrator, 20...casing, 21...porous sintered body, 30...control unit, 100...analysis system, F1...fraction collector, FP...flow path, ME...separation membrane, P1-P3...first to third supply ports, P4...first discharge port, P5...second discharge port, SC...sample container, SP...discharge space, V1...flow rate adjustment valve, V3, V4...switching valve, V5...opening / closing valve, Y...one way, a...first direction, b...second direction, c...target component, p1-p5...piping, p11-p17...piping

Claims

1. The housing and a separation membrane that partitions an internal space of the housing so as to form a flow path within the housing; a first supply unit that supplies the first liquid from a first position in the housing to the flow path so that the first liquid flows along the separation membrane in a first direction; a second supply unit that supplies the second liquid from a second position in the housing to the flow path so that the second liquid flows along the separation membrane in a second direction opposite to the first direction; a third supply unit that supplies a third liquid containing a target component having a size that does not permeate the separation membrane from a third position of the housing to the flow path; The concentrator, wherein the third position is between the first position and the second position in the first direction.

2. 2. The concentrator of claim 1, wherein the third position corresponds to a position where the flow of the first liquid in the first direction and the flow of the second liquid in the second direction collide.

3. The concentrating device of claim 1 or 2, further comprising a first discharge unit that discharges the first, second, and third liquids from a fourth position of the flow path after the second supply unit stops supplying the second liquid and after the third supply unit stops supplying the third liquid.

4. The concentrator of claim 3 , wherein the fourth location is downstream from the second location in the first direction.

5. a control unit that controls the first supply unit, the second supply unit, the third supply unit, and the first discharge unit; The control unit controlling the first supply unit, the second supply unit, and the third supply unit so that, during a first operation, the first liquid flows in the first direction in the flow path, the second liquid flows in the second direction, and the third liquid is supplied to the flow path; 5. The concentrating apparatus according to claim 3 or 4, wherein, during a second operation after the first operation, the first liquid is supplied to the flow path so as to flow in the first direction while the second and third liquids are not supplied to the flow path, and the first, second, and third supply units and the first discharge unit are controlled so that the first liquid is discharged from the flow path together with the second liquid and the third liquid remaining on the separation membrane.

6. During the first operation, the first supply unit supplies the first liquid to the flow path at a first flow rate, and the second supply unit supplies the second liquid to the flow path at a second flow rate greater than the first flow rate; The concentrator of claim 5 , wherein the third position is set closer to the first position in the first direction than the second position.

7. A liquid chromatograph, An analytical system comprising the concentrating device according to any one of claims 1 to 6, which concentrates a specific component in the eluent discharged after analysis by the liquid chromatograph as the target component in the third liquid.

8. The liquid chromatograph a separation column; a mobile phase supply unit that supplies a mobile phase to the separation column; a sample introduction unit that introduces a sample into the mobile phase to be supplied to the separation column by the mobile phase supply unit; a detector for detecting components of the sample in the mobile phase discharged from the separation column; The analytical system according to claim 7 , wherein the third supply unit supplies a mobile phase containing the sample discharged from the detector to the flow path as the third liquid.

9. The analytical system according to claim 8 , wherein the third supply unit supplies one or more components of the sample discharged from the detector as the target component to the flow channel together with the third liquid.

10. supplying a first liquid from a first position in the housing to a flow path along a separation membrane in the housing so that the first liquid flows in a first direction along the separation membrane; supplying a second liquid to the flow path from a second position in the housing so that the second liquid flows along the separation membrane in the housing in a second direction opposite to the first direction; supplying a third liquid containing a target component having a size that does not permeate the separation membrane from a third position in the housing to the flow path; The method of concentrating, wherein the third position is between the first position and the second position in the first direction.

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