JPH01221660A - Extraction chromatograph separator - Google Patents

Abstract

PURPOSE:To improve reproducibility and to widen the range of linearity to sample load by providing selector valves which switch passages passing an extraction vessel, a trap column and sepn. column and passages to bypass the same to each of said vessel and columns. CONSTITUTION:The liquefied gas in a cylinder 1 is fed to a flow passage system by fully opening a pressure regulating valve 21 and operating a pump 3. A 6-way valve 6 is selected to put the supercritical fluid into the extraction vessel 13 to extract the soluble component in a sample. The extract is thereafter passed through the 6-way valves 6, 7 and a pressure regulating valve 14 and is adsorbed by the adsorbent material in the trap column 16. The 6-way valves 6-8 are switched, the pressure regulating valve 14 is fully opened and the pump 4 is operated. The liquefied gas and the modifier solvent pass through a heat exchanger to form the supercritical fluid. The modifier solvent dissolves the extract in the trap column when the 6-way valve 7 is switched at this time. The dissolved extract is passed through the valves 8, 9 and separated by the sepn. column then the components are extracted by a detector 20.

Description

Detailed Description of the Invention [Industrial Application Field] The present invention extracts soluble substances from a sample using a supercritical fluid or liquefied gas, and conducts this extract online to a chromatograph to separate it into components. The present invention also relates to an extraction/chromatographic separation device that can separate the components as needed.

[Prior Art] The present inventor has developed an extraction/chromatography system that is capable of extracting soluble substances from a sample using a supercritical fluid or liquefied gas, and leading this extract online to a chromatograph to separate it into components. Devised a graph separation device (Japanese Patent Application Laid-Open No. 62-276
Publication No. 60).

This extraction and chromatographic separation device injects a supercritical fluid or liquefied gas into an extraction container containing a sample.
The eluting fluid is pumped to extract the soluble components of the sample, and the extract in the flow path, mainly on the downstream side of the extraction container, is stored in a trap filled with the gaseous extraction/elution fluid in advance. The extract was temporarily held in the loop by low-pressure suction, and then led online to the chromatograph.

In addition, on the downstream side of the detector that makes up the chromatograph,
A backpressure control valve was provided to separate the extract into components in the separation column and direct them to the detector.

This device is revolutionary in that extraction and separation can be performed online.

[Problems to be solved by the invention]However, when a sample is extracted, separated into components, and quantified,
The relative standard deviation was about lθ%, the reproducibility was low (and the range of linearity with respect to sample loading was narrow), so it was not suitable for use as a constant fluorescence analyzer.

Furthermore, since a trapping loop was used for sample extraction, only the first sample extracted could be trapped, and the range of sample amounts that could be handled was extremely narrow.

SUMMARY OF THE INVENTION In view of the above problems, an object of the present invention is to provide an extraction/chromatographic separation device and method that can improve reproducibility and widen the range of linearity in the negative direction of the sample and the range of sample amounts that can be handled. It is in.

[Means for Solving the Problems] In the present invention, an extraction section, an adsorption section, and a separation section are sequentially connected in cascade, and an extraction/elution fluid containing a supercritical fluid or liquefied gas is supplied to the extraction section. In the chromatographic separation device, the extraction section includes an extraction container in which a sample is stored, a switching valve that switches the extraction container to a flow path and a passage that bypasses the extraction container, and the extraction section includes comprises a pressure regulating valve, a trap column connected to the downstream side of the pressure regulating valve and filled with an adsorbent for adsorbing extract, a flow path passing through the trap column via the pressure regulating valve, and The separation unit includes a separation column that separates the extract into components, and a switching valve that switches between a flow path passing through the separation column and a flow path bypassing the separation column. The present invention is characterized in that it includes a valve, a detector provided on the downstream side of the switching valve to detect eluate, and a pressure regulating valve connected to the downstream side of the switching valve.

Furthermore, the extraction/chromatographic separation method according to the present invention includes:
This is also a method of using the above device, in which an extraction/elution fluid containing a supercritical fluid or liquefied gas is passed through the sample in an extraction container to extract the soluble components of the sample, and the soluble components are transferred to an adsorbent packed in a trap column. an extraction/adsorption step in which the extract is adsorbed onto an adsorbent by introducing the extract and reducing its solubility by reducing the pressure of the extraction/elution fluid, and then bypassing the trap column and increasing the pressure to reduce its solubility. The increased extraction/elution fluid is guided to the trap column, and by increasing the extraction/elution fluid, the extract adsorbed on the adsorbent is eluted from the trap column, and most of the extract other than the target extract is removed. , bypassing and draining the separation column;
A separation step of passing the target extract through a separation column and bypassing the extraction container and the trap column to pass the extraction/elution fluid through the separation column to separate the target extract into components. It is characterized by

[Example] (1) One example An embodiment of the present invention will be described based on the drawings.

FIG. 1 is a block diagram of one embodiment of the present invention, showing an extraction/chromatographic separation apparatus.

In the figure, l is a cylinder, which contains a liquefied gas, such as liquefied CO, as an extraction/elution fluid.

2 is a storage tank, which contains a modifier solvent, such as ethanol. This modifier solvent forms part of the extraction and elution fluid.

Pumps 3 and 4 pump the liquefied gas at a predetermined flow rate and feed the modifier solvent at a constant ratio to the liquefied gas. This pump 3 is cooled by a cooler (not shown).

5 is a heat exchanger, one end of which is commonly connected to the outlets of pumps 3 and 4, and preheats the liquefied gas into a supercritical fluid.

6 to 9 are hexagonal valves, all of which have boats A to F,
In the illustrated state of deafness, the flow path between the solid line portions AB, CD, and EF is open, and the flow path between the dotted line portions BC, DE, and FA is open. In either case, it is possible to switch between an open channel and an open channel by a switching operation. Boat A of the six-way valve 6 is connected to the other end of the heat exchanger 5. Six-way valve 6-9
Between, boat F of six-way valve 6 and boat A of six-way valve 7 are connected, boat F of six-way valve 7 and boat A of six-way valve 8 are connected, and boat B of six-way valve 8 and boat B of six-way valve 9 are connected. is connected. Further, each boat D of the six-way valves 6 to 8 is closed.

lO~12 is an exhaust valve, and each population is a six-way valve 6
~8 boats C, each outlet is connected to the outside, and the extraction container 13, trap column 16,
When the separation column is separated from the flow path system, that is, when the dotted line portions of the hexagonal valves 6 to 9 are opened, the gas is diffused to the outside.

Reference numeral 13 denotes an extraction container, which houses a sample, and has an inlet connected to boat B of the six-way valve 6, and an outlet connected to boat E of the six-way valve 6. This sample may be either a liquid or a solid (including an analyte); in the case of a liquid, the sample is absorbed into a liquid-absorbing substance and then stored in the extraction container 13.

Reference numeral 14 designates a pressure regulating valve with a small internal volume (for example, 20 μC or less), which is composed of, for example, an electromagnetic valve whose valve stem is driven in the axial direction by a solenoid against the biasing force of a return spring to fully open or fully close the flow path. The inlet is connected to the boat B of the six-way valve 7.

15 is a pressure indicating regulator, which detects the pressure on the upstream side of the pressure regulating valve 14 and controls the pressure regulating valve 1 so that this becomes the set value.
4 and indicates the detected pressure.

When the pressure regulating valve 14 is the above-mentioned electromagnetic valve, for example, it is fully opened when the set pressure is exceeded, and fully closed when the set pressure is lower than the set pressure. In this case, the on/off cycle of the solenoid valve is usually 1 to 20 Hz, and there is no particular problem. Further, the sound pressure may be controlled by adjusting the duty ratio between opening and closing of the solenoid valve. By repeating this opening and closing operation, the actual amount of fluid retained within the solenoid valve becomes extremely small, and it is possible to prevent the extract from retaining within the valve (see Japanese Patent Application No. 62-008652).

16 is a trap column, the inlet of which is connected to the pressure regulating valve 1.
4, and the outlet is connected to the boat E of the hexagonal valve 7, and the interior thereof is filled with an adsorbent, such as fine particles of porous silica gel, that adsorbs the extract extracted from the sample.

17 is a standard sample loop, one end of which is connected to the boat C of the six-way valve 9, and the other end connected to the boat F of the six-way valve 9, and a microsyringe (not shown) is used from the injection port I8 of the six-way valve 9. Standard sample loop! A standard sample solution is injected into 7. Ports D and E of the six-way valve 9 are communicated with the outside, and after the standard sample loop 17 is used, the gas in the loop is diffused to the outside.

19 is a separation column, the inlet of which is connected to port A of six-way valve 9, the outlet connected to port E of six-way valve 8,
It is filled with a stationary phase for separating the sample into its components, such as microparticles of porous silica gel.

20 is a detector, for example a multi-wavelength UV detector, the inlet of which is connected to port F of the six-way valve 8.

Reference numeral 21 denotes a pressure control valve, and when configured to allow fractionation, a pressure control valve with a small internal volume is used like 14, and its inlet is connected to the outlet of the detector 20.

A pressure indicating regulator 22 detects the pressure on the upstream side of the pressure regulating valve 21, adjusts the opening degree of the two pressure regulating valves so that this becomes the set pressure, and indicates the detected pressure.

23 is a fraction collector, which is used to separate the eluate discharged through the pressure regulating valve 2.

24 is a constant temperature bath, the above-mentioned components 1g 5.6 to 9.13
,! 6.17 and 19 are housed and kept at a temperature above a predetermined supercritical temperature, for example, when the liquefied gas is CO
3, the temperature is maintained at a predetermined value 10 to 80°C higher than room temperature.

Next, the operation of this embodiment configured as described above will be explained. First, extraction and adsorption processing is performed, followed by separation and preparative processing.

(A) Extraction/adsorption processing The six-way valves 6 and 8 are opened to the flow paths shown by dotted lines, and the six-way valve 7 is opened.
opens the channel shown by the solid line.

The temperature chamber 24 and pressure indication adjustment @15 are set to values that result in the liquefied gas being a supercritical fluid preferred for extracting the target substance from the sample. Further, the set pressure of the pressure indicating regulator 22 is set to zero, that is, the pressure regulating valve 2I is fully opened.

In the second state, the pump 3 is operated to send the liquefied gas in the cylinder 1 to the channel system. When this liquefied gas passes through the heat exchanger 5, it becomes a supercritical fluid.

Next, the sample is placed in the extraction container 13, and the hexagonal valve 6 is switched to open the flow path shown by the solid line. As a result, the supercritical fluid enters the extraction vessel 13 and extracts the soluble components of the sample. Thereafter, when it passes between EF of the six-way valve 6 and between AB of the six-way valve 7, and passes through the pressure regulating valve 14, the pressure is reduced and its solubility decreases rapidly. Therefore, all of the extract is adsorbed by the adsorbent in the trap column 16. The supercritical fluid becomes a gas and passes through the trap column 16 to the E of the six-way valve 7.
F, the AP of the six-way valve 8, the detector 20, and the pressure regulating valve 21, and are emitted into the atmosphere.

(B) Separation/Preparation Process When the extraction/adsorption process described above is completed, the six-way valves 6 to 8 are switched, and the six-way valves 6 and 7 open the channels shown by dotted lines.
The six-way valve 8 opens the flow path shown by the solid line. Further, the hexagonal valve 9 leaves the flow path shown by the solid line open.

Next, the pressure setting value of the pressure indicating regulator 15 is set to zero, that is, the pressure F4 regulating valve 14 is fully opened, and the pressure setting value of the pressure indicating regulator 22 is set to the pressure that maintains the supercritical fluid on the upstream side. do. In this state, the pump 4 is also operated.

The set flow of liquefied gas and modifier solvent passes through the heat exchanger 5, and this liquefied gas passes through the heat exchanger 5 and becomes a supercritical fluid. between AP of 7, between AB of six-way valve 8, between BA of six-way valve 9,
After passing through the separation column 19, the EF section of the six-way valve 8, and the detector 20, and then through the pressure regulating valve 21, the pressure is reduced to atmospheric pressure, and the supercritical fluid is gasified and released into the atmosphere.

Additionally, the modifier solvent is discarded.

After this state has passed for a certain period of time and the separation column 19 is stabilized, the six-way valve 7 is switched to open the flow path shown by the solid line. In this flow path system, the supercritical fluid and the modifier solvent pass through the trap column 16 to dissolve and separate the extract adsorbed on the adsorbent, and between AB of the six-way valve 8 and BA of the six-way valve 9. It passes through the separation column 19 and is separated into components, and then passes through the FE of the six-way valve 8 to the detector 20.
The component is detected through.

Depending on the case, the target component is fractionated using the fraction collector 23 while being monitored using the detector 20 .

(C) Comparison with a standard sample When analyzing a standard sample and comparing it with the analysis results of the target extract, the following processing is performed.

The six-way valves 6 and 7 open the flow path shown in dotted lines, and the six-way valves 8 and 9 open the flow path shown in solid lines.

In this state, pumps 3 and 4 are activated to pass the supercritical fluid and modifier solvent through the separation column 19. Next, after injecting the standard solution from the injection part 18 into the standard sample loop 17 using the microsyringe, the six-way valve 9 is switched to open the flow path shown by the dotted line. As a result, the standard sample in the standard sample loop 17 is analyzed.

(2) Test example Next, a test example showing the effects of the above device will be explained.

The conditions of the apparatus are as follows.

Extraction column 13: inner diameter 1ha, length 13.8mm Trap column 16: inner diameter 4.6m11. The adsorbent is a porous silica gel separation column 19 with a diameter of 5 μm: inner diameter 4; 6 m 11. length 250m
5 The stationary phase is a polyporous silica gel with a diameter of 5 μm.Pressure regulating valves 14 and 21: These are the above electromagnetic valves, and the valve stem has a diameter of 3
．． 48m haze, stroke length 01mm Detector 20: Ultraviolet/visible multi-wavelength detector multi-
330 type, JASCO Corporation piping: inner diameter 1/16 inch, outer diameter 0.
The conditions for the 5'ms mobile phase are as follows.

Liquefied gas: CO2 flow rate 5.0ag/-in Pressure 200kg/am to make supercritical fluid, temperature 40°C Modifier solvent: Methanol flow rate 0.2 Karasu Q/in As a sample, extraction efficiency error from polymer In order to eliminate the
Irganox 1076) and Irg, 1010 nomethanol solutions were used, and the respective concentrations were adjusted to I B/mρ. Dispense 10μ of this mixed solvent with a microsyringe,
Injected into the extraction container 13, the above extraction, adsorption, separation,
Preparative processing was performed. However, in the separation process, after the separation column 19 was stabilized, the six-way valve 7 continuously opened the flow path shown by the dotted line. The results obtained are shown in Table 1 below.

Table 1 As can be seen from the above table, relative standard deviation (R3D) 1.
Good values of 2 to 1.7% were obtained.

Then know these additives in polypropylene! ! 1
The same operation as above was performed. The set meal results were the same as Soxhlet/HPLC using chloroform. A good reproducibility of R2O3.0% or less was obtained. Furthermore, linearity with respect to sample negative force was also confirmed up to 60 mg. (3) Expansion Note that the present invention includes various other modifications.

For example, extraction or chromatographic separation 9 may be performed using liquefied gas instead of supercritical fluid.

In addition, before extracting the main target extract, extract the non-main target extract under different temperature and pressure conditions from the target extraction, and use the six-way valve 7.8. It may be discharged to the outside through the dotted line channel. In this case, the desired extract can be extracted at a high concentration.

Further, depending on the sample, it is possible to perform extraction and chromatographic separation using only a supercritical fluid, so a modifier solvent is not necessarily necessary.

Furthermore, the flow path switching may be performed in the same manner as described above using another valve instead of the hexagonal valve.

Furthermore, without disposing the detector 20 along the flow path, the eluate may be sequentially fractionated by a plurality of fraction collectors 23 and then analyzed by the detector online or offline.

Furthermore, although a case has been described in which a solenoid is used as the drive device for the valve stem, for example, a piezoelectric element may be used as the drive source, or the valve stem itself may be composed of a piezoelectric element.

This valve may be one that opens and closes a flow path between an inlet and an outlet, and is not limited to one in which the valve stem reciprocates in its axial direction, and may be of a slide spool type, a rotary spool type, or the like.

[Effects of the Invention] As explained above, according to the extraction/chromatographic separation apparatus and method according to the present invention, reproducibility can be sufficiently improved compared to the conventional apparatus, and the linearity with respect to sample load can be improved by 10% compared to the conventional apparatus.
It has the excellent effect of being able to more than double the amount of sample that can be handled, and making the range of sample amounts that can be handled much wider than before.

[Brief explanation of the drawing]

FIG. 1 is a fluid circuit diagram of an extraction/chromatographic separation apparatus according to an embodiment of the present invention. 1: Liquefied gas cylinder 2: Modifier solvent storage tank 3.4: Pump 5: Heat exchanger 6-9; Method valve to-12; Exhaust valve 13: Extraction vessel 14.21: Pressure adjustment valve 15: Pressure indication adjustment with alarm Instrument 16: Trap column 17: Standard sample loop 18: Standard sample injection section 19: Separation column 20: Detector 22: Pressure indicator regulator 23: Fraction collector 24: Constant temperature bath

Claims (1)

[Claims] 1) An extraction/chromatographic separation device in which an extraction section, an adsorption section, and a separation section are sequentially connected in cascade, and an extraction/elution fluid containing a supercritical fluid or liquefied gas is supplied to the extraction section. , the extraction unit includes: an extraction container (13) in which a sample is stored; and a switching valve (6) that switches between a flow path passing through the extraction container (13) and a flow path bypassing the extraction container (13). The adsorption unit includes: a pressure adjustment valve (14); a trap column (16) connected to the downstream side of the pressure adjustment valve (14) and filled with an adsorbent that adsorbs the extract; The trap column (16) via the pressure regulating valve (14)
The separation section includes a separation column (19) that separates the extract into components, and a separation column (19) that separates the extract into components. a switching valve (8) that switches between a flow path passing through the separation column and a flow path that bypasses the separation column; a detector (20) that is provided on the downstream side of the switching valve (8) and detects eluate; The pressure regulating valve (
21) An extraction/chromatographic separation device characterized by having the following. 2) Extraction/elution fluid containing supercritical fluid or liquefied gas is passed through the sample in the extraction container (13) to extract the soluble components of the sample, and the soluble components are extracted to the adsorbent packed in the trap column (16). an extraction/adsorption step in which the extract is adsorbed onto an adsorbent by reducing the pressure of the extraction/elution fluid to reduce its solubility;
), the extraction/elution fluid whose solubility has been increased by pressurization is guided to the trap column, and the extract adsorbed on the adsorbent is eluted from the trap column (16) by increasing the extraction/elution fluid; Most of the extract other than the target extract is discharged by bypassing the separation column (19), and the target extract is discharged through the separation column (19).
) and bypassing the extraction container (13) and the trap column (16) to pass the extraction/elution fluid through a separation column (19) to separate the target extract into components; An extraction/chromatographic separation method characterized by: