PREPARATIVE CHROMATOGRAPH

Abstract

A preparative chromatograph for collecting target components in a sample temporally separated in a column of a chromatograph in respective preparative containers is provided with: a detection unit having a flow cell accommodated in a housing and a detector for detecting a component that passes through the flow cell; a first pipe that connects the column and an inlet end of the flow cell; a flow path switching unit accommodated in the housing and configured to selectively flow the components that passed through the flow cell through a preparative flow path that is a flow path connected to the preparative containers or a waste liquid flow path; and a second pipe accommodated in the housing and connecting an outlet end of the flow cell and the flow path switching unit.

Claims

1. A preparative chromatograph for collecting target components in a sample temporally separated in a column of a chromatograph in respective preparative containers, comprising: a) a detection unit having a flow cell accommodated in a housing and a detector for detecting components that pass through the flow cell; b) a first pipe that connects the column and an inlet end of the flow cell; c) a flow path switching unit accommodated in the housing and configured to selectively flow the components that passed through the flow cell to a preparative flow path which is a flow path to be connected to the respective preparative containers or a waste liquid flow path; and d) a second pipe accommodated in the housing, the second pipe connecting an outlet end of the flow cell and the flow path switching unit.

2. The preparative chromatograph as recited in claim 1, wherein the detection unit is an absorptiometer equipped with an LED light source.

3. The preparative chromatograph as recited in claim 1, wherein the detection unit is equipped with an optical fiber that transmits irradiation light emitted from a light source arranged outside the housing and irradiated to the flow cell.

4. The preparative chromatograph as recited in claim 1, further comprising: e) a fractionation head attached to an outlet end of the preparative flow path, wherein the flow cell, the second pipe, and the flow path switching unit are mounted on the fractionation head; and f) a drive mechanism configured to move the outlet end of the preparative flow path among the preparative containers.

5. The preparative chromatograph as recited in claim 2, further comprising: e) a fractionation head attached to an outlet end of the preparative flow path, wherein the flow cell, the second pipe, and the flow path switching unit are mounted on the fractionation head; and f) a drive mechanism configured to move the outlet end of the preparative flow path among the preparative containers.

6. The preparative chromatograph as recited in claim 3, further comprising: e) a fractionation head attached to an outlet end of the preparative flow path, wherein the flow cell, the second pipe, and the flow path switching unit are mounted on the fractionation head; and f) a drive mechanism configured to move the outlet end of the preparative flow path among the preparative containers.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0026] FIG. 1 is a configuration diagram of a main part of one example of a preparative chromatograph according to an embodiment of the present invention.

[0027] FIG. 2 is an explanatory diagram relating to a fractionation head of the preparative chromatograph of this example.

[0028] FIG. 3 is a configuration diagram of a main part of an absorptiometer of the preparative chromatograph of this example.

[0029] FIG. 4 is a comparison of pipes, etc., between the preparative chromatograph of this example and a conventional preparative chromatograph.

DETAILED DESCRIPTION

[0030] Examples of a preparative chromatograph according to exemplary embodiments of the present invention will be described below with reference to drawings.

[0031] FIG. 1 shows a configuration of a main portion of a preparative chromatograph of this example. FIG. 2 shows a configuration of a main portion of a fraction collector 20 of the preparative chromatograph. The preparative liquid chromatograph of this example is roughly composed of a liquid chromatograph unit 10 for separating target components contained in a sample, a fraction collector 20 for collecting the target components separated by the liquid chromatograph unit 10, and a control unit 30 for controlling operations of these devices.

[0032] In the liquid chromatograph unit 10, a mobile phase in a mobile phase container 11 is sucked up by a liquid supply pump 12 and fed to a column 14 at a predetermined flow rate. A sample containing the target component is injected at a sample injection unit 13 and transported to the column 14 by the flow of the mobile phase. The target components in the sample are temporally separated and eluted within the column 14. The units of the liquid chromatograph unit 10 are accommodated in respective housings and connected by a pipe, respectively.

[0033] The fraction collector 20 is provided with an absorptiometer 21 equipped with three LEDs 211a, 211b, and 211c which are different in light emission wavelength as light sources, a fractionation head 22, a moving mechanism (a rail 23, motor, etc.) of the fractionation head 22, and an electromagnetic valve 24. The absorptiometer 21 and the electromagnetic valve 24 are connected by a second pipe 27 and are accommodated inside of the fractionation head 22 and configured to be moved along the rail 23 together with the fractionation head 22. In the fraction collector 20, a plurality of preparative containers 26 accommodated in a rack 25 are placed. The parts of the fraction collector 20 are accommodated in a single housing 20a.

[0034] The components separated in the column 14 of the liquid chromatograph unit 10 are introduced into the flow cell 212 of the absorptiometer 21 through the first pipe 15. The configuration of the main part of the absorptiometer 21 is shown in FIG. 3. In the absorptiometer 21, three LEDs 211a, 211b, and 211c are light sources that emit light of wavelength bands to be absorbed by three kinds of target components to be prepared, and the light is irradiated in a time division manner (that is, the light emitted from the three LEDs is sequentially irradiated) to the flow cell 212 in accordance with the control signal from a preparative control unit 32. Then, the measurement light that passed through the flow cell 212 is detected by a first photodiode 213. A part of the light emitted from each of the LEDs 211a, 211b, and 211c is detected by a second photodiode 214. The detection signals from the first photodiode 213 and the second photodiode 214 are sent to the control unit 30. In the control unit 30, after calculating the absorbance of light of three kinds of wavelengths, a chromatogram is created and displayed on a screen of the display unit 50, which will be described later.

[0035] When the target component is detected in the absorptiometer 21, the electromagnetic valve 24 is switched by a preparative control unit 32 which will be described later, and the target component that passed through the flow cell 212 is collected into a preparative container through a preparative flow path. After the target component has passed, the electromagnetic valve 24 is switched again by the preparative control unit 32, so that the component that passed through the flow cell 212 is guided to a waste liquid flow path.

[0036] The control unit 30 is equipped with a storage unit 31 and a preparative control unit 32. The preparative control unit 32 is a functional block for controlling the operation of each part of the liquid chromatograph unit 10 and the fraction collector 20. Also, the input unit 40 and the display unit 50 are connected to the control unit 30.

[0037] The preparative control unit 32 makes the display unit 50 display a preparative condition input screen on the display unit 50 so that a user can input a pipe capacity of the second pipe 27 and a liquid supply flow rate of the liquid supply pump 12. When these are input, the delay time is calculated from the pipe capacity and the liquid transfer flow rate and stored in the storage unit 31. The delay time is a time required for the (target) component detected in the absorptiometer 21 to reach the electromagnetic valve 24. The preparative control unit 32 switches the flow path of the electromagnetic valve 24 to the preparative flow path side at the time when the delay time has elapsed from the detection starting point of time of the target component in the absorptiometer 21 to start collection of the target component, and switches the flow path of the electromagnetic valve 24 to the waste liquid flow path side when the delay time has elapsed from the detection end point of time of the target component to finish collection of the target component.

[0038] As described above, the absorptiometer 21 in this example uses the LEDs 211a, 211b, and 211c that each emits light having a wavelength to be absorbed by a target component as light sources. Therefore, it is unnecessary to provide a spectroscope like a conventional absorptiometer using a white light source such as a mercury lamp. Therefore, the absorptiometer 21 is compact and can be accommodated in the fractionation head 22. In addition, in this example, since the internal electromagnetic valve 24 of the fractionation head 22 is also accommodated, the pipe length of the second pipe 27 that connects the flow cell 212 of the absorptiometer 21 and the electromagnetic valve 24 is shorter than the conventional one. Therefore, it is possible to reliably collect a target component by reducing the error of the delay time caused by variation in diameter of the pipe occurring at the time of manufacturing. Furthermore, it is possible to directly connect the flow cell 212 of the absorptiometer 21 and the electromagnetic valve 24 (in this case, the second pipe according to the present invention is a boundary thereof), which enables collection of a target component without delay time. Although FIG. 2 shows an example in which the absorptiometer 21 and the electromagnetic valve 24 are accommodated in the fractionation head 22, the absorptiometer 21 and the electromagnetic valve 24 can be mounted on the upper surface or the side surface of the preparative head 22.

[0039] Regarding each of the configuration of the aforementioned example and a conventionally used configuration (comparative example), hereinafter, the results of calculating the delay time which depends on the pipe capacity from the detector to the electromagnetic valve and the diffusion capacity which depends on the pipe capacity from the column to the electromagnetic valve will be explained. FIG. 4 compares the configuration of this example and that of the comparative example. In both the example and the comparative example, the flow rate was set to 1,000 L/min.

[0040] As shown in FIG. 4, the diameter of the first pipe 15 (the column 14 to the flow cell 212) of the preparative chromatograph of this example was 0.1 mm, the length thereof was 1,000 mm, and the capacity thereof was 7.9 L. The diameter of the second pipe 27 (the flow cell 212 to the electromagnetic valve 24) was 0.1 mm, the length thereof was 50 mm, and the capacity thereof was 0.4 L. On the other hand, in a conventional preparative chromatograph, the diameter of the first pipe (the column to the flow cell) was 0.1 mm, the length thereof was 300 mm, and the capacity thereof was 2.4 L. The diameter of the second pipe (the flow cell to the electromagnetic valve) was 0.3 mm, the length thereof was 1,000 mm, and the capacity thereof was 70.7 L. The diameter of the second pipe is different from the others (thicker than the other) because the pressure resistance of the detector flow cell is low. In other words, if a narrow and long pipe is connected to the outlet end of the flow cell of the detector, the backpressure becomes too high, which causes leakage. On the other hand, in the preparative chromatograph of this example, since the second pipe 27 is short, even if the pipe diameter is small, there is no worry that an excessive back pressure will be applied to the flow cell.

[0041] Under the above conditions, a delay time was obtained by dividing the capacity of the second pipe by the flow rate. As a result, the delay time was 4.24 sec in the comparative example, whereas the delay time was 0.024 sec in this example. That is, it can be understood that the target component can be assuredly collected without substantial delay time.

[0042] For each of the example and comparative example, the diffusion capacity from the column to the electromagnetic valve was determined by the following equation described in Non-Patent Document 1.

[00001]$\begin{array}{cc}\left[\mathrm{Formula}.Math..Math.1\right].Math.& \\ {}_v^2=\frac{.Math..Math.d^4.Math.{\mathrm{LF}}_v}{385.Math.D_m}& \left(1\right)\end{array}$

[0043] In the above formula (1), .sub.v is a diffusion capacity (L), d is a pipe diameter (mm), L is a pipe length (mm), F.sub.v is a flow rate (L/sec), and D.sub.m is a diffusion coefficient (0.002 mm.sup.2/sec, a general value).

[0044] As a specific example, a case is considered in which a target component that passed through a column with a spread corresponding to a peak of 1.0 sec (full width at half maximum). From the above flow rate F.sub.v=1,000 L/min, when the full width at half maximum of the peak of the target component is represented by a flow rate, it becomes 16.67 L. Since a spread of a target component is usually expressed by a Gaussian distribution and, in the Gaussian distribution, the full width at half maximum=2.35 .sub.v, the diffusion capacity of the target component is .sub.v=7.09 L.

[0045] Next, for each of the example and the comparative example, .sub.v is calculated from the aforementioned equation (1) using the pipe diameter and the pipe length shown in FIG. 4, the flow rate F.sub.v=1,000 L/min, and the diffusion coefficient D.sub.m=0.002 mm.sup.2/sec. Then, it becomes .sub.v=2.61 L in the first pipe of this example, and it becomes .sub.v=0.58 L in the second pipe thereof. In the first pipe of the comparative example, it becomes .sub.v=1.43 L, and in the second pipe thereof, it becomes .sub.v=23.46 L. Finally, when the total .sub.v is calculated from the three squared values .sub.v (.sub.v at the time of exiting the column, .sub.v of the first pipe, and .sub.v of the second pipe) by the root mean square, it becomes .sub.v=7.58 L in this example, and it becomes .sub.v=24.55 L in the comparative example. When these values are converted to seconds in accordance with the flow rate F.sub.v, it becomes .sub.v=0.45 sec in this example, and it becomes .sub.v=1.47 sec in the comparative example. Finally, when these are converted to a full width at half maximum, it becomes 1.07 sec in this example and 3.46 sec in the comparative example. That is, in the comparative example, the target component diffuses to the peak of 3.46 sec (full width at half maximum), whereas in this example it is suppressed to the peak of 1.07 sec (full width at half maximum). Therefore, in the preparative chromatograph of this example, the target component can be reliably collected without diffusing the target component in the mobile phase.

[0046] The aforementioned example is merely one example and can be appropriately changed in accordance with the spirit of the present invention. For example, in the aforementioned example, the light from the three types of LEDs 211a, 211b, and 211c is irradiated on the flow cell 212 in a time division manner in the absorptiometer 21. However, in cases where the elution order of the target components that absorb light of each wavelength is known beforehand, the LEDs may be used by switching in that order. Further, the number of LEDs to be used may be appropriately changed. Also, a mercury lamp which is narrow in spectrum like an LED may be used instead of the LED.

[0047] In the aforementioned example, the absorptiometer 21 is used as a detector, but other detectors (a fluorescence detector, an electric conductivity detector, a differential refractive index detector, etc.) can also be used. Also, a plurality of detectors can be used in combination.

[0048] Further, in the same manner as in a conventional detector, an absorption spectrophotometer which uses white light source can also be used. In that case, only the flow cell is placed in the fractionation head of the fraction collector, and a spectroscopic unit (e.g., diffraction grating) for extracting monochromatic light from white light emitted from the light source is placed at an arbitrary position inside or outside the housing of the fraction collector. Then, monochromatic light taken out in the spectroscopic unit can be transported by an optical fiber and irradiated to the flow cell.

[0049] In addition, in the aforementioned example, the absorptiometer 21 is placed inside of the fractionation head 22, but it may be placed at another position within the housing of the fraction collector.

DESCRIPTION OF REFERENCE SYMBOLS

[0050] 10: liquid chromatograph unit [0051] 11: mobile phase container [0052] 12: liquid supply pump [0053] 13: sample injection unit [0054] 14: column [0055] 15: first pipe [0056] 20: fraction collector [0057] 20a: housing [0058] 21: flow cell [0059] 21: absorptiometer [0060] 211a to 211c: LED [0061] 212: flow cell [0062] 213: first photodiode [0063] 214: second photodiode [0064] 22: fractionation head [0065] 23: rail [0066] 24: electromagnetic valve [0067] 25: rack [0068] 26: preparative container [0069] 27: second pipe [0070] 30: control unit [0071] 31: storage unit [0072] 32: preparative control unit [0073] 40: input unit [0074] 50: display unit