JP4955342B2 - Liquid chromatograph apparatus and sample introduction apparatus - Google Patents

Description

  The present invention relates to a liquid chromatograph apparatus and a sample introduction apparatus for a liquid chromatograph apparatus suitable for analysis under high pressure.

  In a liquid chromatograph apparatus, a mobile phase is sucked by a pump unit and is sent to a column together with a sample introduced by a sample introduction unit. The sample introduced into the column is separated into each component and detected by various detectors.

  In general, in an apparatus field called high-performance liquid chromatograph (HPLC), it is required to perform analysis under a high-pressure channel of 20 to 40 MPa at maximum. Such a high-performance liquid chromatograph pump unit is required to be able to supply a mobile phase accurately and precisely even under high pressure.

  Patent Documents 1 and 2 disclose a pump device capable of stable liquid feeding without pulsating flow by controlling the movement of the plunger based on the pressure value detected by the pressure detecting means.

  On the other hand, in the sample introduction unit for high-performance liquid chromatograph, an analysis in which the needle for sucking the sample, the sampling pipe for holding the sample sucked by the needle, and the flow path connecting the injection port are fed at high pressure by the pump unit. A system (direct injection system) incorporated in the flow channel is the mainstream.

  This direct injection method has excellent features in that the sucked sample can be introduced into the column without waste and the carry-over can be reduced because the inside of the needle is always flushed with a mobile phase.

  Patent Document 3 discloses a sample introduction unit in which the inside of a metering pump that sucks a sample through a needle is washed by a mobile phase that is fed by a pump unit.

  In general, in a sample introduction device that employs the direct injection method, the needle can maintain liquid tightness at all times with the seal in the injection port in order to avoid the ejection of the mobile phase from the injection port even in analysis under high pressure. It has a mechanism. During analysis, the needle and sampling pipe incorporated in the high-pressure channel are separated from the high-pressure channel by the channel switching valve when the sample is aspirated, and the solvent in the needle and sampling pipe is released to atmospheric pressure. .

  Then, after the pressure is released to atmospheric pressure, the needle is inserted into the sample holding container, the sample is sucked into the needle or the sampling pipe, and then the needle moves to the above-described injection port. At this stage, the flow path switching valve is switched again, and the sample is supplied to the column together with the mobile phase fed by the pump unit.

Japanese Patent No. 2564588 Japanese Patent No. 2604362 Japanese Patent No. 2730951

  However, in the conventional sample introduction unit, the needle opened to the atmospheric pressure and the solvent in the sampling pipe are pressurized to the system pressure (pressure during analysis) by the pump unit after the sample is sucked into the needle or the like. , Will be supplied to the column.

  That is, a certain time is required until the entire volume in the needle and the sampling pipe is pressurized, and the system pressure becomes unstable during this time. Since this pressure fluctuation appears as a baseline fluctuation on the chromatogram, it overlaps with the peak of the component having a fast retention time (a component that is weakly retained on the column), which causes fluctuations in the retention time and area value of the peak. .

  The peak retention time is a basic qualitative information that is specific to the component separated by the liquid chromatograph, and the area value represents the concentration of the component separated by the liquid chromatograph. The accompanying baseline variation results in inaccurate qualitative information on the chromatogram, which impairs the accuracy and reproducibility of quantitative measurements.

  In particular, when a differential refractive index detector that detects the refractive index difference of a solvent or an absorbance detector that employs a single beam photometry method is used, the influence of pressure fluctuations on the detection value is extremely large due to the principle of the detector. Therefore, such fluctuations in system pressure are naturally undesirable.

  Further, when it is desired to introduce a large-capacity sample into the flow path, it is necessary to increase the internal volume of the sampling pipe, so that the volume of the solvent that is pressurized to the system pressure increases, resulting in a large pressure fluctuation.

  Furthermore, in the apparatus field called ultra high performance liquid chromatograph, since it is required to perform analysis under a pressure of 60 to 100 MPa at the maximum, the baseline fluctuation accompanying the pressure fluctuation at the time of sample introduction becomes more remarkable.

  An object of the present invention is to realize a liquid chromatograph apparatus and a sample introduction apparatus for a liquid chromatograph apparatus that can reduce baseline fluctuation due to pressure fluctuation at the time of sample introduction.

  The sample introduction apparatus according to the present invention introduces a sample from a sample introduction flow path into a chromatograph apparatus having an analysis flow path.

  The sample introduction apparatus according to the present invention includes a sample suction and pressurizing unit that sucks a sample and pressurizes the sample introduction channel, a switching unit that switches separation and connection between the analysis channel and the sample introduction channel, and a sample Control means for controlling the operation of the suction and pressurizing means and the switching means, the control means separates the analysis channel and the sample introduction channel from each other by the switching means, and opens the sample introduction channel to atmospheric pressure, The sample is sucked by the sample suction and pressurization means, the inside of the sample introduction flow path is sealed and pressurized, and the sample introduction flow path and the analysis flow path are connected to each other by the switching means.

  The chromatographic apparatus according to the present invention includes a pump for introducing a mobile phase, a separation column to which a mobile phase and a sample introduced by the pump are supplied, and a detector for detecting the sample by supplying the sample from the separation column. And a sample introduction channel for introducing a sample.

  The chromatographic apparatus according to the present invention includes a sample suction and pressurizing unit that sucks a sample and pressurizes the sample introduction channel, a switching unit that switches separation and connection between the analysis channel and the sample introduction channel, and a switching The analysis channel and the sample introduction channel are separated from each other by means, the sample introduction channel is opened to atmospheric pressure, the sample is sucked by the sample suction and pressurization unit, and the inside of the sample introduction channel is sealed and pressurized. And a control means for connecting the sample introduction flow path and the analysis flow path to each other by the switching means.

  In addition, a sample introduction method according to the present invention includes a pump for introducing a mobile phase, a separation column to which a mobile phase and a sample introduced by the pump are supplied, and a detector for supplying the sample from the separation column and detecting the sample. A sample is introduced from a sample introduction channel into a chromatograph apparatus having an analysis channel having

  The sample introduction method of the present invention separates the analysis channel and the sample introduction channel from each other, opens the sample introduction channel to atmospheric pressure, sucks the sample into the sample introduction channel, and introduces the sample. The inside of the channel is sealed and pressurized, and the sample introduction channel and the analysis channel are connected to each other.

  A liquid chromatograph apparatus, a sample introduction apparatus for a liquid chromatograph apparatus, and a sample introduction method that can reduce baseline fluctuation due to pressure fluctuation during sample introduction can be realized.

  Embodiments of the present invention will be described below with reference to the accompanying drawings.

  FIG. 1 is a schematic configuration diagram of a liquid chromatograph apparatus according to a first embodiment of the present invention.

  In FIG. 1, the mobile phase 1 sent by the pump unit 2 is sent to the column 3 via the flow path switching valve 6, the sampling pipe 22, the needle 11, the injection port 10, the pipe 16, and the flow path switching valve 6. .

  On the other hand, the sample introduced by the needle 11 is sent to the column 3 together with the mobile phase 1 and detected by the detector 4. The detector 4 is connected to the drain 5.

  Here, the system pressure applied to the analysis flow path composed of the pump 2, the column 3, and the detector 4 is detected by a pressure detection means (not shown) installed in the pump unit 2, and is maximum in the high-performance liquid chromatograph apparatus. 20 to 40 MPa, reaching a maximum of 60 to 100 MPa in an ultrahigh performance liquid chromatograph.

  The flow path switching valve 6 has six ports composed of P1 to P6, and performs two-position switching by rotating 60 degrees. Further, the flow path switching valve 7 has six ports composed of P1 to P6. Then, the port of the flow path switching valve 7 rotates to 3 positions by 45 degrees, so that the pipe 23 connected to the port P6 is connected to no port (blocked), port P2, and port P3, respectively. .

  Operation control of the pump 2, the flow path switching valves 6 and 7, the needle 11, the plunger 12, which will be described later, and the like is performed by an operation control unit (not shown).

  As a preparatory step before entering the sample aspirating step, first, the high-pressure flow path constituted by the sampling pipe 22, the needle 11, the injection port 10, and the pipe 16 is separated from the analysis flow path and opened to the atmospheric pressure.

FIG. 2 is a flow chart of the pressure release process for separating the high pressure flow path from the analysis flow path. In FIG. 2, the flow path switching valve 7 is rotated 45 degrees counterclockwise from the state shown in FIG. 1, and the flow path switching valve 6 is rotated 60 degrees counterclockwise, so that the high pressure flow path is The pressure separated from the passage and loaded on the solvent in the sampling pipe 22, the needle 11, the injection port 10, and the pipe 16 is released to atmospheric pressure via the pipe 18, the pipe 21, the cleaning port 9, and the drain 20. . At this time, the pipe 16 communicates with the pump chamber 8 via the switching valve 6, the pipe 17, the switching valve 7, and the pipe 23.

  Next, the sample suction step will be described. FIG. 3 is a flow chart of the sample suction process. In FIG. 3, the flow path switching valve 7 is rotated 45 degrees counterclockwise from the state shown in FIG. 2, and the needle 11 is pulled out from the injection port 10 and moved to the sample holding container 24 holding the sample 15. . At this time, the sampling pipe 22 communicates with the pump chamber 8 via the switching valves 6 and 7.

  In this state, the aspirated sample is held in the needle 11 and the sampling pipe 22 by retracting the plunger 12 constituting the sample suction and pressurizing means. Before and after the sample suction step, the needle 11 is immersed in the cleaning port 9 as necessary to clean the outer wall of the needle 11 (not shown).

  Next, a pressurization process for repressurizing the flow path whose pressure has been temporarily reduced to atmospheric pressure by the sample suction process will be described. FIG. 4 is an explanatory diagram of a pressurization process for repressurizing the flow path whose pressure has been reduced to atmospheric pressure.

  In FIG. 4, after the sample is aspirated, the needle 11 moves to the injection port 10 and the liquid tightness is maintained. In this state, the plunger 12 is advanced to compress the solvent in the flow path, so that the flow path constituted by the sampling pipe 22, the needle 11, the injection port 10, and the pipe 16 opened to the atmospheric pressure rises to the system pressure. Let

In the pressurizing step, the distance dx for moving the plunger 12 forward is obtained as follows, for example. When the volume change with respect to the pressure dP is dV, the isothermal compressibility k _T (constant) of the solvent having the volume V is expressed by the following equation (1).

k _T = (− dV / V) / dP ――― (1)
The pump unit 2 is provided with a pressure sensor for detecting the pressure in the analysis flow path. The required dP is determined from the system pressure P detected by the pressure sensor, and the sample introduction flow path (high pressure flow path). The volume V of the solvent to be compressed is determined from the physical dimensions of the elements constituting the above and the current position of the plunger 12.

Isothermal compressibility k _T is the solvent-specific constants, since the diameter of the plunger 12 is known, dV, i.e. dx is determined. Further, the sample solvent in substantially atmospheric pressure, mobile phase, considering that consists of a plurality of kinds of cleaning liquid, by adding the temperature-dependent coefficient compressibility k _T, dx can be determined more accurately.

  Thereby, the difference between the system pressure (pressure in the analysis channel) and the pressure in the sample introduction channel after the pressurization step is reduced, and the baseline fluctuation on the chromatogram can be reduced.

  After the pressurizing step, the flow path switching valve 6 is rotated 60 degrees clockwise from the state shown in FIG. 4, whereby the sampling pipe 22, the needle 11, the injection port 10, and the pipe 16 are incorporated as the analysis flow path. A flow chart at this time is shown in FIG.

  In FIG. 5, the sample held in the needle 11 and the sampling pipe 22 is supplied to the column 3 together with the mobile phase 1 by the pump unit 22. After rotating the flow path switching valve 6, the plunger 12 is advanced in a state where the flow path switching valve 7 is rotated 45 degrees clockwise from the state shown in FIG. , 21 to the cleaning port 9.

  Further, in order to sufficiently replace the solvent in the washing port 9, the flow path switching valve 7 is further rotated 45 degrees clockwise (the state shown in FIG. 1) and the plunger 12 is moved backward as necessary. The cleaning liquid 14 is sucked into the pump chamber 8. Thereafter, the flow path switching valve 7 is further rotated 45 degrees counterclockwise (returns to the state shown in FIG. 5), and the cleaning liquid 14 supplied from the switching valve 7 and the pipe 19 to the pump chamber 8 is discharged to the cleaning port 9. To do. In addition, 13 is a seal | sticker and 25 is a spring.

  As described above, according to the first embodiment of the present invention, the analysis flow path and the high pressure flow path are separated, the high pressure flow path is opened to atmospheric pressure, and the plunger 12 is moved to enter the pump chamber 8. By sucking the solvent, the sample is sucked from the needle 11 into the high-pressure channel. Then, after sealing the high-pressure channel from atmospheric pressure, the plunger 12 is moved to supply the solvent in the pump chamber 8 to the high-pressure channel, and the pressure in the high-pressure channel is made substantially equal to the pressure in the analysis channel. Thereafter, the high-pressure channel and the analysis channel are connected, and the sample is supplied to the detector 4 via the column 3.

  Therefore, it is possible to realize a liquid chromatograph that can reduce baseline fluctuations due to pressure fluctuations during sample introduction.

  In addition, a sample introduction device having a high-pressure channel separate from the analysis channel, that is, the switching valves 6 and 7, the sampling pipe 22, the needle 11, the injection port 10, the pump chamber 8, the plunger 12, and an operation control unit (not shown). It is possible to realize a sample introduction device that includes the pipes 16 to 21 and can reduce baseline fluctuation due to pressure fluctuation during sample introduction.

  Furthermore, it is possible to realize a sample introduction method capable of reducing baseline fluctuation due to pressure fluctuation during sample introduction.

  Next, a second embodiment of the present invention will be described. FIG. 6 is a schematic configuration diagram of a liquid chromatograph apparatus according to the second embodiment of the present invention.

  The difference between the second embodiment and the first embodiment of the present invention is that the pressure sensor 26 is installed in the pipe 19 part. Since other configurations are the same as those of the first embodiment and the second embodiment, description thereof will be omitted. The pressure sensor 26 is connected to the switching valve 7 via a pipe 27 and communicates with the pump chamber 8 via a pipe 28.

  The pressure sensor 26 detects the pressure in the flow path accompanying the movement of the plunger 12 in the pressurizing step, and a signal corresponding to the detected pressure is supplied to a drive motor (not shown) that drives the plunger 12. The drive motor adjusts the position of the plunger 12 in accordance with a detection signal from the pressure sensor 26.

  That is, the system pressure (pressure in the analysis channel) is detected by a pressure sensor arranged in the pump unit 2, and the control unit (not shown) detects the detected system pressure and the high-pressure channel (sample introduction channel). The position of the plunger 12 is controlled so that the pressure in the parenthesis becomes equal. Thereby, since the pressure fluctuation when the sampling pipe 22, the needle 11, the injection port 10, and the pipe 16 are incorporated in the analysis flow path can be suppressed as much as possible, the baseline fluctuation at the time of sample introduction can be reduced.

  Here, the pressure sensor 26 does not need to be disposed at the position of FIG. 6, and may be disposed anywhere in the flow path from the port P5 (blocked) to the port P2 (blocked) of the flow path switching valve 7.

  As a typical pressure sensor of the present invention, a semiconductor strain gauge pressure transducer is used in which pressure is converted into displacement through a diaphragm made of a stainless member having excellent corrosion resistance and taken out as an electric quantity by a bridge circuit.

  This sensor has high robustness, reliability and stability even when used under high pressure for a long time, has excellent corrosion resistance to various solvents such as organic solvents, acids, alkalis, etc. It is suitable as a liquid chromatograph because of its small size, small solvent contact capacity (dead volume) of the detection unit, excellent response, and use in a wide pressure range.

  Also in the second embodiment of the present invention, as in the first embodiment, a liquid chromatograph apparatus, a sample introduction apparatus for a liquid chromatograph apparatus, and a sample introduction capable of reducing baseline fluctuation due to pressure fluctuation at the time of sample introduction A method can be realized.

  Next, the results of obtaining chromatograms using the liquid chromatograph apparatus when the present invention is applied and when it is not applied will be compared.

  FIG. 7 is a chromatogram obtained when the present invention is not applied. FIG. 7A shows the result when a blank is introduced, and FIG. 7B shows the result when a sample is introduced.

  When the present invention is not applied, since a large baseline fluctuation occurs when a blank is introduced, the target component peak and the baseline fluctuation overlap when the sample is introduced. Since the baseline fluctuation waveform accompanying the pressure fluctuation at the time of sample introduction is generally poor in reproducibility, it is extremely difficult to separate the component peak waveform and the baseline fluctuation waveform by waveform processing calculation after measurement. is there.

  Therefore, when the present invention is not applied, a large error may occur when calculating the peak area, and correct quantification is performed for such a peak that is eluted quickly from the column (fast retention time). Difficult to get results. In addition, if the retention time of the target component peak is earlier and it completely overlaps with the baseline fluctuation due to pressure fluctuation, there is a risk that the peak retention time, which is qualitative information, may be erroneously recognized, and the data The result is unreliable.

  FIG. 8 is a chromatogram obtained when the present invention is applied. By realizing a system in which the atmospheric pressure channel that sucks the sample is pressurized to the system pressure and incorporated into the analysis channel, pressure fluctuations and baseline fluctuations can be extremely reduced when a blank is introduced (Fig. 8 (A)). As a result, as shown in FIG. 8B, it is possible to obtain accurate retention times and area values with good reproducibility even for peaks that are eluted quickly from the column (fast retention times). Therefore, the reliability of the analysis result can be improved.

  In addition, although the example mentioned above is an example at the time of applying this invention to a liquid chromatograph apparatus, this invention is applicable also to a gas chromatograph apparatus and a supercritical chromatograph apparatus.

It is a schematic block diagram of the 1st Embodiment of this invention. It is a flow path figure showing the pressure relief process in a 1st embodiment of the present invention. It is a channel figure showing a sample suction process in a 1st embodiment of the present invention. It is a channel figure showing a pressurization process in a 1st embodiment of the present invention. It is a channel figure showing a washing process in a 1st embodiment of the present invention. It is a schematic block diagram of the 2nd Embodiment of this invention. It is a figure which shows the chromatogram when this invention is not applied. It is a figure which shows the chromatogram at the time of this invention being applied.

Explanation of symbols

  DESCRIPTION OF SYMBOLS 1 ... Mobile phase, 2 ... Pump unit, 3 ... Column, 4 ... Detector, 5 ... Drain, 6, 7 ... Flow path switching valve, 8 ... Pump chamber , 9 ... Cleaning port, 10 ... Injection port, 11 ... Needle, 12 ... Plunger, 13 ... Seal, 14 ... Cleaning liquid, 15 ... Sample, 16-18 ... Pipe, 19, 20 ... drain, 21, 22, 23, 27, 28 ... pipe, 24 ... sample holding container, 25 ... spring

Claims (3)

In a sample introduction apparatus for introducing a sample from a sample introduction flow path into a chromatograph apparatus provided with an analysis flow path,
Sample suction and pressurizing means for sucking the sample and pressurizing the sample introduction channel;
Switching means for switching separation and connection between the analysis channel and the sample introduction channel;
A first pressure sensor for detecting the pressure in the analysis flow path;
A second pressure sensor for detecting the pressure in the sample introduction channel;
Control means for controlling the operation of the sample suction and pressure means and the switching means;
And the control means separates the analysis flow path and the sample introduction flow path from each other by the switching means, opens the sample introduction flow path to atmospheric pressure, and the sample introduction flow by the sample suction and pressurization means. After the sample is sucked into the channel, the inside of the sample introduction channel is sealed, and the pressure value in the sample introduction channel detected by the second pressure sensor is detected by the first pressure sensor. Further, the inside of the sample introduction flow path is pressurized by the sample suction and pressurization means so that the pressure value in the analysis flow path becomes equal, and the sample introduction flow path and the analysis flow path are connected to each other by the switching means. A sample introduction device characterized by that. An analysis flow path having a pump for introducing a mobile phase, a separation column to which a mobile phase and a sample introduced by the pump are supplied, a detector for supplying the sample from the separation column and detecting the sample, and a sample In a chromatograph apparatus provided with a sample introduction channel to be introduced,
Sample suction and pressurizing means for sucking the sample and pressurizing the sample introduction channel;
Switching means for switching separation and connection between the analysis channel and the sample introduction channel;
A first pressure sensor for detecting the pressure in the analysis flow path;
A second pressure sensor for detecting the pressure in the sample introduction channel;
The analysis channel and the sample introduction channel are separated from each other by the switching unit, the sample introduction channel is opened to atmospheric pressure, and the sample is sucked into the sample introduction channel by the sample suction and pressurization unit. After that, the inside of the sample introduction channel is sealed, and the pressure value in the sample introduction channel detected by the second pressure sensor is the pressure value in the analysis channel detected by the first pressure sensor. Control means for pressurizing the inside of the sample introduction flow path by the sample suction and pressurization means, and connecting the sample introduction flow path and the analysis flow path to each other by the switching means;
A chromatographic apparatus comprising: A chromatograph comprising an analysis flow path having a pump for introducing a mobile phase, a separation column to which a mobile phase and a sample introduced by the pump are supplied, and a detector for supplying the sample from the separation column and detecting the sample In a sample introduction method for introducing a sample into a device from a sample introduction channel,
The analysis channel and the sample introduction channel are separated from each other,
The sample introduction channel is opened to atmospheric pressure, and a sample is sucked into the sample introduction channel by one sample suction and pressurization means ,
A first pressure sensor for sealing the inside of the sample introduction channel and detecting the pressure in the analysis channel detects the pressure in the analysis channel and detects the pressure in the sample introduction channel . the second pressure sensor detects the pressure of the sample introduction flow path, the pressure value of the sample introduction flow path is such that the pressure value of the analysis passage, the sample introduction channel in said one Pressurize by means of sample suction and pressurization,
A sample introduction method comprising connecting the sample introduction channel and the analysis channel to each other.

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