JP2016008908A - Component extracting and separating device, and component extracting and separating method, using supercritical fluid - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a component extracting and separating device and a component extracting and separating method, both using supercritical fluid, that make available spectra reduced in baseline fluctuations attributable to pressure fluctuations.SOLUTION: A component extracting and separating device comprises a channel 111 that passes a mobile phase and a sample to a column 11; a regulator 14 that adjusts pressure applied on the mobile phase in the channel 111 to a pressure surpassing a critical pressure, which turns the mobile phase in the column 11 into a supercritical state; a column oven 12 for heating the column 11 to a temperature surpassing a critical temperature; a cooler 130 that cools the mobile phase having passed the column 11; and a detector 13 that detects components of the sample in the mobile phase cooled by the cooler 130.

Description

  The present invention relates to a component extraction / separation apparatus and a component extraction / separation method for extracting a target component from a sample using a supercritical fluid. The present invention also relates to a chromatograph using a supercritical fluid.

  Supercritical Fluid Chromatograph (SFC) is a chromatograph using a fluid (supercritical fluid) having a temperature and pressure exceeding a critical point (critical temperature, critical pressure) as a mobile phase. It has the characteristic that the melt | dissolution power of a mobile phase can be controlled by adjusting. In particular, a supercritical fluid chromatograph using carbon dioxide as the mobile phase of the supercritical fluid is often used as an analyzer that can analyze in a short time by increasing the flow rate of the mobile phase because the viscosity of the fluid is low. (See Patent Document 1).

  FIG. 4 is a schematic configuration diagram of such a conventional supercritical fluid chromatograph. The supercritical fluid chromatograph 40 includes a column 41, a column oven 42, a detector 43, a back pressure regulator 44, a liquid feed pump 45a, 45b, a mobile phase source 46, an autosampler 47, and a collection unit 48 that houses a plurality of collection containers 48a. The mobile phase source 46 includes a cylinder 46a that supplies a supercritical fluid (for example, carbon dioxide having a critical pressure of 7.4 MPa and a critical temperature of 31 ° C.) and a solvent container 46b that supplies a solvent such as methanol. At one end of the column 41, an input side channel 411 connected to the liquid feed pumps 45a and 45b is provided, and an autosampler 47 is provided in the middle of the input side channel 411. Further, an output side flow path 412 connected to the detector 43 is provided at the other end of the column 41, and a back pressure regulator 44 is provided at the end. As the detector 43, a multi-wavelength detector using a PDA (Photo Diode Array), a UV detector, or the like is used.

  The supercritical fluid chromatograph 40 operates as follows. First, the back pressure regulator 44 sets the pressure of the mobile phase in the input side channel 411, the column 41 and the output side channel 412 between the liquid feed pumps 45a and 45b and the back pressure regulator 44 to a pressure exceeding the critical pressure. Adjust to. Although it depends on the type of supercritical fluid used, in general, the pressure is increased to about 10 MPa. Next, the liquid feed pumps 45 a and 45 b flow the mobile phase (supercritical fluid and solvent) from the mobile phase source 46 to the column 41 through the input side channel 411. The sample is injected into the mobile phase by the autosampler 47, flows through the input side flow path 411 by the mobile phase, and is carried to the column 41.

  The column 41 is disposed in the column oven 42 and is adjusted to a temperature (for example, 40 ° C.) exceeding the critical temperature of the mobile phase (supercritical fluid) to be used. Therefore, the mobile phase passing through the column 41 is in a supercritical state here. The sample that has entered the column 41 from the input side channel 411 interacts with the stationary phase of the column 41 and is separated for each component, and sequentially comes out from the output side channel 412 at different times for each component. Here, the time at which the separated components come out can be known in advance by measuring a standard sample having known components.

  For accurate detection, it is important to detect quickly without changing the state of the components of the sample coming out of the column 41. Therefore, the components of the sample separated as described above are detected by the detector 43 in the supercritical state near the outlet of the column 41. The detection data detected by the detector 43 is sent to a data processing device such as a computer (not shown). The data processing device creates a spectrum for each component from the detection data. The separated components can be quantified from the shape of the spectrum. After the sample separated for each component is detected by the detector 43, it passes through the back pressure regulator 44 and is collected in a collection container 48a installed in the collection unit 48 together with the mobile phase.

JP 2010-243215 A

  In the supercritical fluid chromatograph 40, in order to maintain the inside of the flow path in a supercritical state, the back pressure regulator 44 controls the pressure in the flow path to be kept constant. However, in practice, it is difficult to keep the pressure in the flow path constant, and a pulsation of about 0.01 MPa occurs. It is known that the density of a supercritical fluid changes greatly with a slight change in pressure (see Patent Document 1). This change in density causes the baseline to fluctuate in the spectrum of the component detected by the detector 43.

  The present invention has been made in view of the above points, and an object of the present invention is to provide a component extraction / separation apparatus and component extraction using a supercritical fluid capable of obtaining a spectrum with a small baseline fluctuation. It is to provide a separation method.

The component extraction / separation apparatus according to the present invention, which has been made to solve the above problems,
a) a flow path for the mobile phase and sample through the column;
b) a regulator for adjusting the pressure applied to the mobile phase in the flow path to a pressure exceeding the critical pressure, which is a pressure at which the mobile phase becomes supercritical;
c) a column oven for heating the column to a temperature above the critical temperature, which is the temperature at which the mobile phase in the column is in the supercritical state;
d) a cooling unit for cooling the mobile phase that has passed through the column;
e) a detector for detecting a component of the sample in the mobile phase cooled by the cooling unit;
It is characterized by providing.

In addition, the component extraction and separation method according to the present invention, which has been made to solve the above problems,
a) applying a pressure exceeding the critical pressure, which is a pressure at which the mobile phase becomes supercritical, to the mobile phase in the channel through which the mobile phase and the sample pass through the column;
b) heating the column to a temperature above the critical temperature, which is the temperature at which the mobile phase in the column is in the supercritical state;
c) detecting the component of the sample after cooling the mobile phase that has passed through the column by a cooling unit;
It is characterized by providing.

  The cooling unit can cool the mobile phase to a temperature below the critical temperature at which the mobile phase becomes liquid. Such a cooling unit may use a Peltier element, or may be an air cooling device or a water cooling device. Moreover, it may be attached to the detector and integrated with the detector.

  As a result of conducting various experiments while changing the conditions of each part of the supercritical fluid chromatograph, the present inventor has found that the pressure changes as the mobile phase containing the sample components, which is the object to be detected by the detector, is cooled. It was experimentally found that the fluctuation of the baseline with respect to was reduced, and the present invention was achieved. This result is contrary to the conventional idea that rapid detection without changing the state of the sample components is important for accurate detection, but rather changes the state of the sample components coming out of the column. This is surprising because it enables accurate detection with small baseline fluctuations.

  According to the component extraction / separation apparatus and component extraction / separation method of the present invention having the above-described configuration, the detector detects the sample components after the mobile phase that has passed through the column is cooled by the cooling unit. It is possible to obtain a spectrum in which the fluctuation of the baseline due to the pressure pulsation of a certain mobile phase is small. Thereby, it is possible to obtain a measurement result with high reliability of the quantitative result and high repeatability.

BRIEF DESCRIPTION OF THE DRAWINGS The schematic block diagram of the supercritical fluid chromatograph which is one embodiment of this invention. The schematic block diagram of the detector which is one embodiment of this invention. The figure which shows the extent of the fluctuation | variation in the baseline of a spectrum with respect to the change of a pressure about the case where the temperature of a mobile phase is (a) critical temperature (31 degreeC), (b) 35 degreeC, (c) 40 degreeC. . The schematic block diagram of the conventional supercritical fluid chromatograph.

  FIG. 1 is a schematic configuration diagram of a supercritical fluid chromatograph which is an embodiment of a component extraction / separation apparatus according to the present invention. A supercritical fluid chromatograph 10 includes a column 11, a column oven 12, a detector 13, A back pressure regulator 14, liquid feed pumps 15 a and 15 b, a mobile phase source 16, an autosampler 17, and a recovery unit 18 that houses a plurality of recovery containers 18 a are provided. The mobile phase source 16 includes a cylinder 16a that supplies a supercritical fluid and a solvent container 16b that supplies a solvent such as methanol. At one end of the column 11, an input side flow path 111 connected to the liquid feed pumps 15 a and 15 b is provided, and an autosampler 17 is provided in the middle of the input side flow path 111. Further, an output-side flow path 112 connected to the detector 13 is provided at the other end of the column 11, and a back pressure regulator 14 is provided at the end.

  The supercritical fluid chromatograph 10 has a cooling unit 130 attached to the detector 13 and is configured to cool the mobile phase flowing through the output side flow path 112. Different. In the present embodiment, carbon dioxide is used as the supercritical fluid, and a UV detector is used as the detector 13. As shown in FIG. 2, the detector 13 includes a flow cell 131, a heat insulating material 132, a lens 133, and a temperature measurement unit 134. The detector 13 is thermally shielded from the outside by the heat insulating material 132, and is hardly affected by external temperature changes. In the present embodiment, the cooling unit 130 has a Peltier element 130 a and a heat radiating fin 130 b and is attached to the heat insulating material 132 so as to be integrated with the detector 13. The lens 133 is a lens through which light incident on the flow cell 131 and light emitted from the flow cell 131 pass, and is disposed adjacent to the flow cell 131 and embedded in the heat insulating material 132. The temperature measuring unit 134 can be a thermocouple having a platinum resistor.

  The supercritical fluid chromatograph 10 operates as follows. First, the back pressure regulator 14 sets the pressure of the mobile phase in the input side flow path 111, the column 11 and the output side flow path 112 between the liquid feed pumps 15a, 15b and the back pressure regulator 14 to a pressure exceeding the critical pressure. Adjust to. In the present embodiment, the pressure is about 10 MPa which is a pressure exceeding the critical pressure (7.4 MPa) of carbon dioxide which is a mobile phase. Next, the liquid feed pumps 15 a and 15 b flow the mobile phase (supercritical fluid and solvent) from the mobile phase source 16 to the column 11 through the input side flow path 111. The sample is injected into the mobile phase by the autosampler 17, flows through the input side flow path 111 by the mobile phase, and is carried to the column 11.

  The column 11 is disposed in a column oven 12 and is adjusted to about 40 ° C., which is a temperature exceeding the critical temperature (31 ° C.) of carbon dioxide as a mobile phase. Therefore, the mobile phase passing through the column 11 is in a supercritical state here. The sample that has entered the column 11 from the input side channel 111 interacts with the stationary phase of the column 11 and is separated for each component, and sequentially comes out from the output side channel 112 at different times for each component. Here, the time at which the separated components come out can be known in advance by measuring a standard sample having known components.

  The mobile phase containing the components of the sample thus separated enters the detector 13 through the output side channel 112 and flows through the coiled channel 113. The mobile phase is cooled by the cooling unit 130 while flowing through the coiled channel 113. A temperature measurement unit 134 provided near the coiled flow channel 113 measures the temperature of the mobile phase flowing through the coiled flow channel 113. The measured temperature is detected by a temperature control circuit 135 provided outside the detector 13, and the temperature control circuit 135 feedback-controls the amount of current supplied to the Peltier element 130 a of the cooling unit 130.

  The mobile phase containing the sample components flows through the flow cell 131 after being cooled as described above. In the flow cell 131, ultraviolet light from a light source (not shown) is irradiated to a mobile phase including a sample component via a lens 133, and the light absorption characteristic that the sample component absorbs the ultraviolet light is measured. Since the light absorption characteristic varies depending on the substance, the component of the sample in the flow cell can be specified by such optical detection.

  The sample component detection data in the flow cell 131 is sent to a data processing device such as a computer (not shown). The data processing device creates a spectrum for each component from the detection data. The separated components can be quantified from the shape of the spectrum. The sample separated for each component is detected by the flow cell 131 of the detector 13, and then collected through the back pressure regulator 14 in the collection container 18 a installed in the collection unit 18 together with the mobile phase.

(Example)
Using the supercritical fluid chromatograph 10 described above, the pressure of the mobile phase in the input side flow path 111, the column 11 and the output side flow path 112 between the liquid feed pumps 15a, 15b and the back pressure regulator 14 is changed. In this case, an experiment was conducted to confirm the position of the baseline of the spectrum of a specific sample component detected by the detector 13. Results of experiments conducted when the temperature of the mobile phase was set to the critical temperature (31 ° C.) and 35 ° C. by the cooling unit 130 and when the cooling unit 130 was not operated and the temperature was the same as the column oven 12 (40 ° C.). It shows to Fig.3 (a)-FIG.3 (c). 3A to 3C, the horizontal axis represents the mobile phase pressure, and the vertical axis represents the baseline position.

  It can be seen from FIGS. 3A to 3C that the position of the baseline changes linearly with respect to the pressure. The absolute value of the slope of the linear approximation decreased with decreasing temperature of the mobile phase containing the sample components. From this result, it was found that as the mobile phase containing the components of the sample was cooled, the fluctuation of the baseline with respect to the change in pressure was reduced. This is a tendency obtained in an experiment in which the pressure is intentionally changed, but this is also true for a pulsation of about 0.01 MPa.

  That is, after the mobile phase that has passed through the column is cooled by the cooling unit, the detector 13 detects the sample component, and thus the spectrum with a small baseline fluctuation due to the pressure pulsation of the mobile phase in the supercritical state is small. Can be obtained. Thereby, it is possible to obtain a measurement result with high reliability of the quantitative result and high repeatability.

  Further, as can be seen from FIG. 3A, even when the mobile phase is cooled to the critical temperature (31 ° C.) at which the mobile phase becomes a liquid, the fluctuation of the baseline with respect to the change in pressure is reduced. This is contrary to the conventional idea that it is important for accurate detection to detect quickly without changing the state of the component of the sample, and the state of the component of the sample coming out of the column 11 is determined. Rather, it was a surprising result that changing it enabled accurate detection with little baseline variation.

  It should be noted that the above embodiment is merely an example of the present invention, and it is obvious that modifications, corrections, and additions may be made as appropriate within the scope of the present invention, and included in the claims of the present application. In the above embodiment, the example in which the Peltier element 130a is attached as the cooling unit 130 so as to be integrated with the detector 13 has been described, but the configuration of the component extraction / separation device according to the present invention is not limited thereto. Any structure that can cool the mobile phase containing the sample component flowing in the output-side flow path 112 between the column 11 and the flow cell 131 of the detector 13 may be used. Instead of the Peltier element 130a, other devices such as an air cooling device or a water cooling device may be used. You may have a cooling device. The cooling unit 130 may be a unit separated from the detector 13.

  In the above embodiment, an example in which carbon dioxide is used as the supercritical fluid has been described, but other known supercritical fluids may be used. Moreover, although the example which uses a UV detector as the detector 13 was shown, the multiwavelength detector using a PDA (Photo Diode Array), an optical rotation detector, a circular dichroism detector used in a liquid chromatograph, Any one of a fluorescence detector, a refractive index detector, and an evaporative light scattering detector may be used.

DESCRIPTION OF SYMBOLS 10, 40 ... Supercritical fluid chromatograph 11, 41 ... Column 12, 42 ... Column oven 13, 43 ... Detector 14, 44 ... Back pressure regulator 15a, 15b, 45a, 45b ... Liquid feed pump 16, 46 ... Mobile phase Sources 16a, 46a ... cylinders 16b, 46b ... solvent containers 17, 47 ... autosamplers 18, 48 ... collection units 18a, 48a ... collection containers 111, 411 ... input side channels 112, 412 ... output side channels 113 ... coiled Flow path 130 ... Cooling unit 130a ... Peltier element 130b ... Radiating fin 131 ... Flow cell 132 ... Insulating material 133 ... Lens 134 ... Temperature measuring unit 135 ... Temperature control circuit

Claims (6)

a) a flow path for the mobile phase and sample through the column;
b) a regulator for adjusting the pressure applied to the mobile phase in the flow path to a pressure exceeding the critical pressure, which is a pressure at which the mobile phase becomes supercritical;
c) a column oven for heating the column to a temperature above the critical temperature, which is the temperature at which the mobile phase in the column is in the supercritical state;
d) a cooling unit for cooling the mobile phase that has passed through the column;
e) a detector for detecting a component of the sample in the mobile phase cooled by the cooling unit;
A component extraction / separation apparatus comprising:   The component extraction / separation apparatus according to claim 1, wherein the cooling unit cools the mobile phase to a temperature equal to or lower than the critical temperature at which the mobile phase becomes a liquid.   The component extraction / separation device according to claim 1, wherein the cooling unit is one using a Peltier element, an air cooling device, or a water cooling device. a) applying a pressure exceeding the critical pressure, which is a pressure at which the mobile phase becomes supercritical, to the mobile phase in the channel through which the mobile phase and the sample pass through the column;
b) heating the column to a temperature above the critical temperature, which is the temperature at which the mobile phase in the column is in the supercritical state;
c) detecting the component of the sample after cooling the mobile phase that has passed through the column by a cooling unit;
A component extraction / separation method comprising:   The component extraction / separation method according to claim 4, wherein the cooling unit cools the mobile phase to a temperature equal to or lower than the critical temperature at which the mobile phase becomes a liquid.   The component extraction / separation method according to claim 4, wherein the cooling unit is a unit using a Peltier element, an air cooling device, or a water cooling device.

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