JP4860367B2 - Liquid chromatograph - Google Patents

Description

  The present invention relates to a liquid chromatograph apparatus.

  In a liquid chromatograph, a separation column is used to separate each component of a sample. However, when measuring a biological sample, a large amount of contaminants contained in the sample hinder measurement and hinder high-sensitivity analysis. Not only is the degradation of the separation column a major factor.

  The service life of a separation column due to deterioration depends on the type of measurement sample and the environment in which it is used, so it is difficult to specify it quantitatively, and it is judged based on the deterioration in the quality of actual measurement data and fluctuations in the pressure of liquid delivery. . However, there are a number of factors other than the deterioration of the separation column that cause a decrease in the quality of the measurement data and the fluctuations in the liquid feeding pressure, and the determination is difficult, and there is a current situation that relies heavily on the empirical rules of the measurer.

  Therefore, in Patent Document 1, after identification and quantification, the resolution is calculated using the data of two peaks for resolution calculation designated in advance, and the calculated resolution is compared with a reference value. A technique for determining whether a separation column is normal or deteriorated is described.

JP-A-7-120451

  By the way, the deterioration state of the separation column can be known from fluctuations in the liquid supply pressure, etc., but there are many other causes of fluctuations in the liquid supply pressure besides the deterioration of the separation column, and its accurate judgment is based on many years of experience. Reliable part is big and difficult.

  Further, when a trouble occurs in the liquid chromatograph apparatus, the relationship between the occurrence of the trouble and the introduced sample can be determined if the time of introduction of the sample into the separation column can be determined.

  However, in the conventional technique, when a trouble related to the sample occurs in the liquid chromatograph apparatus, it is not possible to easily determine the time point when the sample is introduced into the separation column.

  In the column switching method in the prior art, the setting of the optimum sample introduction time is complicated and requires a long time.

  Here, the column switching method is a method of automatically concentrating target components and removing impurities when measuring a biological sample. A trap column is connected via a high-pressure switching valve before the separation column. In addition, by switching the flow path of the mobile phase using a high-pressure switching valve, the target component can be automatically concentrated and impurities can be removed.

  The sample is first introduced into the trap column, temporarily held in the trap column, and then introduced into the separation column by switching the mobile phase flow path by switching the high-pressure switching valve. The time required for introducing the sample into the trap column can be calculated from the volume in the flow path and the flow velocity.

  However, since the arrival of the sample is actually delayed due to diffusion in the flow path or the like, the sample introduction time is set longer than the calculated value in view of the likelihood.

  If the sample introduction time is too long, components with weak adsorptive power pass through without being held in the trap column, and even if the high-pressure switching valve is switched, the components do not reach the separation column and the sample can be analyzed. Absent. Conversely, if the sample introduction time is too short, the sample does not reach the trap column, so even if the high-pressure switching valve is switched, the component does not reach the separation column and the sample cannot be analyzed.

  In the column switching method in the prior art, the optimization of the sample introduction time can only be determined by experimental investigation, and this experimental examination requires complicated labor and a long time.

  An object of the present invention relates to the realization of a liquid chromatograph apparatus capable of automatically judging the deterioration of a separation column and the time for introducing a sample.

  The liquid chromatograph apparatus according to the present invention has a visible portion where a part or the whole of a separation column for separating a sample introduced by a sample introduction means is formed of a transparent material and the internal material is visible. A light source that irradiates light to the visible part, a detection part that detects light reflected from the visible part, and a data processing part that determines deterioration of the internal material of the separation column based on the reflected light detected by the detection part With.

  In addition, the liquid chromatograph apparatus of the present invention has a visible portion in which a part or the whole of the trap column that temporarily holds the sample introduced by the sample introduction means is formed of a transparent material and the internal material can be seen. The light source that irradiates light to the visible part of the trap column, the detection part that detects light reflected from the visible part of the trap column, and the internal material of the trap column based on the reflected light detected by the detection part A data processing unit for judging deterioration.

  It is possible to realize a liquid chromatograph apparatus capable of automatically judging the deterioration of the separation column and the sample introduction time.

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 sample introduced from the injector 2 is sent to the separation column 4 by feeding the mobile phase 6 by the feeding pump 1. Then, after the sample is separated by the separation column 4, it is introduced into the sample analyzer 5 and analyzed.

  The separation column 4 is disposed in the column oven 9. In the column oven 9, a light source 7 for ultraviolet light and visible light and a detection unit 8 are arranged. In addition, the separation column 4 includes a visible portion 3 formed of a transparent member that can visually see the inside of the column.

  The light from the light source 7 is irradiated on the visible portion 3, is reflected by the internal material of the separation column 4, and enters the detection portion 8. The detection unit 8 is connected to a data processing unit (not shown) outside the column oven 9.

  While the analysis by the liquid chromatograph apparatus is repeated, the impurities contained in the sample accumulate in the separation column 4, thereby causing deterioration of the separation column 4. A sample derived from a living body contains a large amount of contaminants, and typical examples of such contaminants include lipids, proteins, and carbohydrates.

  It is known that these impurities are colored brown, for example, by Maillard reaction when adsorbed on the separation column 4, and the wavelength of ultraviolet light or visible light reflected by the visible portion 3 is detected by the detecting portion 8. By detecting and supplying the data to the data processing unit, it is possible to digitally monitor the deterioration of the separation column 4. A display unit is connected to the data processing unit, and the degree of deterioration of the separation column 4 can be displayed on the display unit.

  In addition, the data processing unit detects that the sample has been introduced into the separation column 4 by adding a fluorescent label or adding and labeling a standard substance with a fluorescent label in the sample. It is possible to record the time when the sample is introduced into the separation column 4.

  As described above, according to the first embodiment of the present invention, the visible portion 3 through which the internal material can be seen is formed in the separation column 3, and the result detected by the detection portion 8 of the optical system is processed by the data processing portion. In addition, the deterioration of the internal material of the separation column can be automatically determined. Furthermore, since the timing at which the sample is introduced into the separation column 4 can be detected and recorded, when the quality of the measurement data deteriorates or the pressure fluctuates, the above-mentioned deterioration in quality occurs at any time since the introduction of the sample. Can be determined. This can be used to determine the cause of the above-described deterioration in quality.

FIG. 2 is a schematic configuration diagram of a liquid chromatograph apparatus according to the second embodiment of the present invention.
In FIG. 2, the measurement sample is introduced into the trap column 10 from the sample introduction unit 12 and temporarily held. Next, by switching the flow path in the hexagonal valve 11, the sample is sent to the analysis column 4 by the liquid feed pump 1 and fed by the mobile phase 6 and separated. Thereafter, the sample is introduced into the sample analyzer 5 and analyzed.

  The trap column 10 is provided with a visible portion 13 formed of a transparent member that allows the inside of the trap column 10 to be visible. Although not shown, a light source for irradiating the visible part 13 of the trap column 10 with ultraviolet rays or visible light and a detection part are arranged, and the detection part is connected to the data processing part.

  A sample is trapped from the sample introduction unit 12 by adding a fluorescent label to the sample or adding a standard substance to which a fluorescent label is added to the sample, labeling, and monitoring the visible part by the fluorescence detection unit and the data processing unit. When introduced into the column 10, it is possible to measure the time required to reach the trap column 10 from the time when the sample is introduced into the sample introduction unit 12.

  FIG. 3 shows a schematic diagram of a chromatogram obtained by monitoring the trap column 10 in the fluorescence detection unit and the data processing unit. In FIG. 3, the vertical axis indicates the signal intensity, and the horizontal axis indicates the elapsed time. By switching the hexagonal valve 11 when the peak of the signal intensity shows a maximum value, it is possible to send the sample to the analysis column 4 without losing it and realize a highly sensitive analysis.

  Even when the sample has absorption at a specific wavelength of light, the same effect can be obtained by monitoring the visible portion 3 with an ultraviolet-visible absorptiometer.

  Further, as in the first embodiment, it is possible to automatically determine the deterioration inside the trap column 10.

  In the second embodiment, it is possible to form a visible portion in both the separation column 4 and the trap column 10 and monitor the inside.

  In the above-described example, a part of the separation column and the trap column is the visible part, but the whole may be formed of a transparent material.

It is a schematic block diagram of the liquid chromatograph apparatus which is the 1st Embodiment of this invention. It is a schematic block diagram of the liquid chromatograph apparatus which is the 2nd Embodiment of this invention. It is the schematic of a chromatogram when a sample reaches | attains a trap column.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Liquid feed pump 2 Injector 3, 13 Visible part 4 Separation column 5 Sample analysis part 6 Mobile phase 7 Light source 8 Detection part 9 Column oven 10 Trap column 11 Six-way valve 12 Sample introduction part

Claims (4)

In liquid chromatograph equipment,
Sample introduction means;
Liquid feeding means;
A separation column for separating the sample introduced by the sample introduction means;
An analysis unit for analyzing the sample separated by the separation column;
A trap column that is partially or wholly formed of a transparent material, has a visible portion that allows the internal material to be visible, and temporarily holds the sample introduced by the sample introduction means;
A light source for irradiating the visible part of the trap column with light;
A detector that detects light reflected from the visible portion of the trap column;
A data processing unit that determines deterioration of the internal material of the trap column based on the reflected light detected by the detection unit;
A switching valve for switching between a flow path for supplying the sample from the sample introduction means to the trap column and a flow path for supplying the liquid from the liquid supply means to the separation column via the trap column;
A liquid chromatograph apparatus comprising: 2. The liquid chromatograph according to claim 1, wherein the data processing unit stores a time point when the sample reaches the trap column. 2. The liquid chromatograph according to claim 1, wherein the separation column is partially or entirely made of a transparent material, has a visible portion that allows the internal material to be visible, and irradiates the visible portion of the separation column with light. And a detection unit that detects light reflected from the visible part of the separation column, and the data processing unit degrades the internal material of the separation column based on the reflected light detected by the detection unit The liquid chromatograph apparatus characterized by judging. 4. The liquid chromatograph according to claim 3, wherein the data processing unit stores a point in time when the sample reaches the separation column.

Images