JP2014115104A - Analyzer, specimen analytic method, and computer readable storage medium - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a technology for creating a calibration curve capable of easily and efficiently producing a standard specimen appropriate for a measurement concentration zone of a component to be quantitatively analyzed, in a quantitative analysis using the standard specimen containing a plurality of components.SOLUTION: A dilution series of a standard specimen is produced, in a component concentration range from a minimum concentration to a maximum concentration in a measurement concentration zone of a contained component. At this time, the component concentration in the standard specimen is constant for all components. When a calibration curve of each component is created, the calibration curve is created by selectively using only a standard specimen measurement result in the concentration range including the measurement concentration zone of each component. Thus, an accurate calibration curve can be created while the labor for producing the standard specimen is reduced.

Description

  The present invention relates to an analysis apparatus, a sample analysis method, and a computer-readable storage medium. For example, the present invention relates to a calibration curve creation technique created for quantifying an analyte in mass spectrometry.

  In the field of chemical analysis, the measurement intensity of an analytical device may change every several hours, every day, or every analyte depending on the stability of the device itself or external factors. For this reason, when attempting to measure the concentration and content of components (including elements, compounds, etc., hereinafter referred to as representative components) contained in a sample, the user of the analyzer must use a sample with a known concentration at the start of the analysis. , And the relationship with the signal intensity output from the apparatus is obtained. A graph showing this relationship is called a calibration curve. As described above, this calibration curve is necessary because the measurement intensity changes or the measurement results vary from device to device.

  When it is desired to know the concentration of a component contained in an unknown sample, the sample is analyzed by an apparatus, and the concentration corresponding to the output signal intensity may be read from the calibration curve. The calculation that creates a calibration curve and calculates the concentration of the component contained in the sample is called quantitative calculation. When quantitatively calculating the concentration of the target component, it is necessary to measure all the standard samples and unknown samples of the target component under the same conditions so that the signal intensity with respect to the concentration does not change every measurement.

  In recent years, GC / MS (Gas Chromatography / Mass Spectrometry) and LC / MS (Liquid Chromatography / Mass Spectrometry) analysis methods have been used as devices capable of separating and detecting multiple components contained in a single sample. Attention has been paid to the analysis apparatus. In particular, LC / MS analyzers are being applied to the medical field. For example, in therapeutic drug monitoring (TDM) for confirming the drug effect when a drug is administered to a patient, factors related to the therapeutic effect and side effects are determined. Set individualized dosage and administration for each patient while monitoring. Therefore, the administration plan is based on the results of the measurement and analysis of blood concentration and clinical findings. In other words, grasp the blood concentration of the components of multiple drugs, adjust so that a certain component is in a predetermined concentration range, and adjust another concentration range to a different concentration range, or check the range To do. For this purpose, an automatic blood analyzer capable of highly sensitive measurement of blood drug concentration is expected.

  In an automatic analyzer that measures a drug concentration in blood, the type of drug to be measured differs for each sample, that is, a patient. In addition, the concentration of the drug to be measured that is assumed to be present in the sample varies greatly depending on the type of drug. This is because the blood drug concentration (treatment area) where the therapeutic effect appears is different for each drug. As an example, in the case of sodium valproate used as an antiepileptic agent, the therapeutic range is 50-100 μg / mL, but the therapeutic range of tacrolimus used as an immunosuppressant is 2-20 ng / mL, more than 1000 times There is a difference in density. That is, an automatic medical analyzer is required to simultaneously measure various types of drugs in a wide range of concentrations assumed for each drug.

  In addition, when it is desired to accurately quantify the analyte in the LC / MS apparatus, it is common to use the internal standard method. The internal standard method is an analysis method in which a component having physical properties very similar to the component to be quantified is contained in all standard samples at a constant concentration, and the intensity ratio between the signal intensity of this component and the component to be quantified is measured intensity.

  As an internal standard substance, a stable isotope compound of an isotope-labeled analyte or a compound having similar chemical physical properties is generally used. That is, a stable isotope compound and an analog compound whose response to the mass spectrometer is similar to the analysis object and can be measured separately from the analysis object are selected.

  When the analyte and the internal standard substance are simultaneously measured in the mass spectrometer, the non-linearity and peak intensity fluctuations of the two apparatus signal intensities due to the apparatus state exhibit the same behavior. In addition, when a factor such as a decrease in peak intensity or a decrease in ionization efficiency due to a contaminated component occurs, an increase or decrease in the peak area shows the same behavior in the analyte and the internal standard material.

  In other words, in order to accurately create a calibration curve for quantification of an analyte in an LC / MS apparatus, a solution in which a plurality of analytes to be measured and an internal standard substance are mixed with two or more different points. It is necessary to measure by concentration. This requires time and effort to prepare standard samples and analyze those samples. At the same time, human error may occur during preparation and measurement.

  As a specific example, the sodium valproate listed in the previous section has an effective blood concentration of 50 to 100 μg / mL, and when it is desired to accurately quantify the blood concentration range with a four-point calibration curve, for example, The first concentration is 200 μg / mL, which is twice or more of the blood treatment area concentration upper limit, the second concentration is 100 μg / mL, which is the upper limit of the blood treatment area concentration, and the third concentration is the blood treatment. If a calibration curve is created with 4 points of 50 μg / mL, which is the lower limit value of the blood region concentration, and 20 μg / mL, where the fourth concentration is 1/2 times or less the lower limit of the blood treatment region concentration, 50 of the concentration range to be measured ˜100 μg / mL can be accurately quantified.

  Similarly, the therapeutic range for tacrolimus is 2 to 20 ng / mL, and in order to prepare a 4-point calibration curve, for example, by measuring concentrations of 50, 10, 5, 1 ng / mL, etc., a calibration curve is created. 2 to 20 ng / mL of the concentration range to be measured can be accurately quantified.

  As an internal standard substance, for example, 100 μg / ml of ethyl paraoxybenzoate is added to all standard samples. By using the ratio of the measurement intensity of sodium palproate and tacrolimus as the measurement target components and the measurement intensity of ethyl paraoxybenzoate as the internal standard substance as the signal intensity in the calibration curve, the nonlinearity of the instrument signal intensity, It prevents the fluctuation of the peak intensity due to the apparatus state from directly changing the concentration value.

  Therefore, the standard sample takes the above therapeutic range into consideration, the first standard sample is sodium valproate 200 μg / ml, tacrolimus 50 ng / ml and ethyl parahydroxybenzoate 100 μg / ml, and the second standard sample is sodium valproate. 100 μg / ml, tacrolimus 10 ng / ml and paraoxybenzoate 100 μg / ml, the third standard sample is sodium valproate 50 μg / ml, tacrolimus 5 ng / ml and ethyl paraoxybenzoate 100 μg / ml, the fourth standard Four series of standard samples containing 20 μg / ml sodium valproate, 1 ng / ml tacrolimus, and 100 μg / ml ethyl paraoxybenzoate are prepared as samples.

  It is possible to introduce a standard sample containing components prepared at the above concentrations into an LC / MS apparatus and measure three components simultaneously by the component separation function. Moreover, the measurement sensitivity of each component can be individually varied by adjusting the measurement conditions such as the ion permeation amount of the ion mass separation mechanism of the MS section.

Japanese Patent Laid-Open No. 5-79984

  However, in preparing a standard sample, it is necessary to consider a standard sample concentration appropriate for quantifying each drug concentration. This is because the appropriate concentration may vary depending on each drug (component). Therefore, as the number of components to be measured increases, it becomes more difficult to prepare a standard sample. Although the above-mentioned standard reagent preparation example has two components, the clinical analyzer requires a simultaneous analysis performance of 15 to 20 components, and the preparation of the standard sample is more difficult and error-prone.

  As a method for automatically preparing a standard sample, for example, in Patent Document 1, in order to improve the efficiency of analysis, one prepared high concentration calibration sample is used, and automatic dilution is performed as many times as necessary. Repeat the measurement. This reduces the labor and human error of preparing multiple types of standard solutions.

  However, in the method of Patent Document 1, it is necessary for the user to pay attention to the concentration range for measuring each component, and it takes much time to prepare a standard sample. For this reason, it is desirable to be able to reduce the labor of preparing a standard sample for samples containing components of various types and concentration ranges when measuring components in the standard sample and creating a calibration curve.

  The present invention has been made in view of such a situation, and in quantitative analysis using a standard sample containing a plurality of components, it is easy and efficient to prepare a standard sample suitable for the measurement concentration range of the component to be quantified. It is intended to provide a technique for creating a calibration curve that can be carried out automatically.

  In order to solve the above-mentioned problems, the present invention can set whether or not to apply the measurement intensity to the calibration curve for each component contained in the measured standard sample and to set the concentration value for each component for each component. It has become.

  In other words, the analyzer according to the present invention displays a standard sample storage unit for storing a standard sample containing a plurality of components at a predetermined concentration, and a GUI for instructing the dilution rate of the standard sample for each of the plurality of components. In response to an instruction from the control unit that receives the dilution rate of each component indicated by the user and displayed on the screen of the device, the standard sample is diluted based on the dilution rate of each component, A sample introduction unit for introducing a diluted standard sample, a separation unit for separating each component from the diluted standard sample, and obtaining a component having a concentration corresponding to the designated dilution rate, and a designated dilution rate. An analysis unit that analyzes each component of the corresponding concentration and measures the intensity of each component of each concentration. And a control part produces | generates the calibration curve of each component based on the measurement result by an analysis part.

According to the present invention, in a quantitative analysis using a standard sample containing a plurality of components, a standard sample suitable for the measurement concentration range of the component to be quantified can be easily and efficiently produced.
Further features related to the present invention will become apparent from the description of the present specification and the accompanying drawings.

It is a figure which shows the schematic apparatus structure of the analyzer (LC / MS apparatus) by embodiment of this invention. It is a flowchart for demonstrating the analysis process performed in the 1st and 2nd embodiment of this invention. It is a figure which shows the example of the measurement density | concentration area | region setting screen (GUI) in a measurement density | concentration area | region setting process (S103). It is a figure which shows the example of the calibration curve preparation result by the 1st Embodiment of this invention. It is a figure which shows the example of the measurement density | concentration area | region setting screen which enabled it to designate the density | concentration of each component by the 2nd Embodiment of this invention. It is a flowchart for demonstrating the analysis process including the process which confirms and corrects the produced calibration curve by the 3rd Embodiment of this invention. It is a figure which shows the example of the confirmation screen (GUI) whether the calibration curve is produced appropriately. It is an example of a calibration curve after correcting the problem. It is a figure which shows the example of a GUI screen containing an analysis condition setting area | region and a mass calibration area | region.

  Hereinafter, embodiments of the present invention will be described with reference to the accompanying drawings. In the accompanying drawings, functionally identical elements may be denoted by the same numbers. The attached drawings show specific embodiments and implementation examples based on the principle of the present invention, but these are for understanding the present invention and are not intended to limit the present invention. Not used.

  This embodiment has been described in sufficient detail for those skilled in the art to practice the present invention, but other implementations and configurations are possible without departing from the scope and spirit of the technical idea of the present invention. It is necessary to understand that the configuration and structure can be changed and various elements can be replaced. Therefore, the following description should not be interpreted as being limited to this.

  Furthermore, as will be described later, the embodiment of the present invention may be implemented by software running on a general-purpose computer, or may be implemented by dedicated hardware or a combination of software and hardware.

  In the following, each process in the embodiment of the present invention will be described mainly using “device control unit (processor)” as a subject (operation subject). Since the communication port (communication control device) is used, the program may be described as a subject (operation subject). Further, the processing by the program may be processing performed by a computer such as a management server or another information processing apparatus. Part or all of the program may be realized by dedicated hardware, or may be modularized. Various programs may be installed in each computer by a program distribution server or a storage medium.

(1) First Embodiment <Apparatus Configuration>
FIG. 1 is a diagram showing a typical apparatus configuration according to an embodiment of the present invention. Note that the device configuration and embodiment described below are examples for specifically explaining the invention, and the embodiment is not limited to the description. FIG. 1 assumes an LC / MS apparatus.

  The LC / MS apparatus 200 according to the embodiment includes a sample introduction unit 201 that dilutes a standard sample and introduces the sample into the apparatus, a separation unit 202 that separates the introduced sample into each component, and each component obtained by separation. The ionization unit 203 for ionizing, the mass analysis unit 204 for performing mass spectrometry processing on each ionized component, the sample storage unit 205, the eluent storage unit 206, and the respective units are instructed to control the operation. A device control unit 207 that executes a calibration curve calculation process, a display device 208 that outputs a control result by the device control unit 207, and an operation unit 209 that is operated by a user and inputs an instruction. doing.

  The sample introduction unit 201 continuously sucks the eluent 206 in a steady state, and continues to send the solution to the separation unit 202 at a constant pressure. In synchronism with the measurement start timing, the sample 205 is injected into the separation unit by a specified volume. Although the sample 205 is shown as a single sample, it may be an automated apparatus that installs a plurality of samples placed on a tray or the like and sequentially supplies the samples at the start of measurement. The sample introduction unit 201 dilutes the sample based on the dilution concentration (see FIG. 3) designated by the user through the operation unit 209 and introduces the sample into the separation unit 202.

  The separation unit 202 has a portion called a “column” in which a cylindrical container is filled with a filler serving as a stationary phase, and separates the mobile phase sent from the sample introduction unit 201. By changing the packing material in the column, the type of separation action can be changed. For example, if highly polar silica gel is used as the filler, the sample can be separated by the difference in the polarity of the substance.

  The ionization unit 203 is, for example, an electrospray ionization (ESI) or an atmospheric pressure chemical ionization (APCI) method, and can ionize a sample separated by the separation unit. Yes.

  The mass analysis unit 204 includes, for example, an ion mass separation mechanism such as a quadrupole type, an ion trap type, and a time-of-flight type, and a channeltron, a dynode type, or a scintillator and a photodetector that detect separated ions. And a container.

The apparatus control unit 207 controls operations of the sample introduction unit 201, the separation unit 202, the ionization unit 203, and the mass analysis unit 204, stores measurement data obtained by the mass analysis unit 204, and performs arbitrary data processing (calibration curve calculation). Display on the display device 208.
The user operates the operation unit 209 as necessary, and instructs the apparatus control unit 207 to perform an analysis operation.

<Analysis procedure>
FIG. 2 is a flowchart for explaining the most typical analysis procedure according to the embodiment of the present invention. Note that the procedure of FIG. 2 describes only the procedures that are relevant in explaining the contents of the present invention, and procedures that are less relevant in explaining the present invention, such as pretreatment of analytical samples, preparation of the apparatus, and cleaning up. Such a procedure or detailed operation of the apparatus during the measurement operation is omitted.

(I) Analysis condition setting (S102)
When the procedure of analysis start (S101) is advanced, sample analysis condition setting (S102) is first executed. In this step, an analyst (user) uses the operation unit 209 to set analysis conditions, and the apparatus control unit 207 receives the analysis conditions. Settings include the number of samples and measurement order, measurement components, number of standard samples, number of unknown samples, separation unit pressure and temperature, separation time, mass number scanning range in the mass analysis unit, and detector sensitivity settings for each component. is there. The measurement component, the separation time of the separation unit and the mass number scanning range of the mass analysis unit are closely related, and the analyst needs to set with considerable knowledge. In this embodiment, since only the concentration of the target component contained in the sample and the signal intensity when the sample is measured are considered, the measurement conditions are assumed to be set appropriately, and details are omitted. . Information such as measurement conditions is stored in a storage device built in the device control unit 207 as necessary. Alternatively, the measurement conditions stored in advance in the apparatus control unit 207 may be called up and used.

(Ii) Measurement concentration range setting process (S103)
Next, the apparatus control unit 207 executes a measurement density area setting process (S103). An example of a typical measurement concentration range setting screen for performing this step is shown in FIG. In FIG. 3, a measured concentration range setting screen 301 is a graphical user interface (GUI) that is displayed on the display unit 208 and allows the user to make various instructions using the operation unit 209 as necessary. This screen is assumed to be an application that runs on, for example, Windows (registered trademark), which is a computer OS made by Microsoft Corporation. This screen is in a table format, and the concentration of the standard sample before dilution and the dilution series (six types of standard samples 1 to 6 in the example in the figure) are arranged in the horizontal direction. The number of dilution series can be arbitrarily changed on another screen. In the vertical direction, the dilution ratio is arranged in the uppermost row, and the names of the components are arranged in the second and subsequent rows.

  In the present embodiment, in the measurement concentration region setting process (S103), the analyzer applies the measurement intensity to the calibration curve for each component included in the standard sample to be measured using the measurement concentration region setting screen as shown in FIG. Whether or not, and the density value when applied can be set for each component. In the case of measuring by the internal standard method, the internal standard component is measured in addition to the components specified in FIG. 3, but the illustration is omitted in this embodiment. The shaded portion in the table of FIG. 3 is a portion where the value is automatically set by the apparatus control unit 207 and cannot be changed by the analyst.

  The standard sample pre-dilution concentration 302 is a location where the concentration of each component included in a high-concentration standard sample prepared in advance is set. In the conventional quantitative analysis method, the concentration of each component had to be adjusted in consideration of the concentration range to be measured. However, in the method of the present invention, the simplest example is as shown in FIG. , All may be of equal concentration.

  As for the setting of the standard sample dilution ratio 304 of the standard samples 1 to 6, the dilution ratio is input by the analyst when the standard sample dilution series preparation process (S104) is executed. When a numerical value is input to this item, the apparatus control unit 207 instantaneously calculates the concentration of each component in the standard sample and reflects it in the table. For example, since the concentration of theophylline is 200 μg / ml and the dilution ratio of standard sample 1 is 1: 0.025, the concentration of theophylline in standard sample 1 is 5 μg / ml. The dilution ratio at the end of the dilution series (sixth in the example of FIG. 3) is fixed at 1: 1 and is not actually diluted.

  To the right of each component concentration value, an analyzer can give a concentration adoption mark 305. For the measurement values of the concentration / component whose concentration adoption mark 305 is not checked, the intensity measurement value is not adopted when the calibration curve is created. That is, in the calibration curve creation step 108, for each component, a calibration curve is created using only the measured intensity value of the standard sample having the concentration adoption mark 305.

  In the measurement concentration range setting process (S103), the analyst uses the screen of FIG. 3 to determine the standard sample concentration that is the highest concentration among all the components necessary for creating a calibration curve before dilution of the standard sample. The density is set, and the apparatus control unit 207 receives them. The therapeutic range of theophylline, which is a drug used for the treatment of respiratory diseases, is 8 to 15 μg / ml, and the therapeutic range of sodium palproate is 50 to 100 μg / ml. There should be 200 μg / ml, which is about twice the upper limit of the therapeutic range for sodium.

  Next, the apparatus control unit 207 determines the dilution ratio of the dilution series for creating a calibration curve. From the above-mentioned treatment range, for example, 5 μg / ml, which is about a half of the lower treatment range lower limit of theophylline, is set as the first concentration of the standard sample dilution series. The ratios of the second to fifth concentrations may be set so as to include about 1 to 2 points of the therapeutic area of each component. In the example of FIG. 3, the dilution ratio is set so that the second to fifth concentrations are 10, 20, 50, and 100 μg / ml. Finally, a concentration adoption mark 305 is set in accordance with the concentration range (an assumed unknown sample concentration range) to be measured for each component. Since the therapeutic range of theophylline is 8-15 μg / ml, a calibration curve suitable for quantifying the concentration in the therapeutic range can be obtained by preparing a standard curve with standard sample concentrations of 5, 10, 20 μg / ml. Therefore, a check is given to 10, 20, 50, and 100 μg / ml of the concentration adoption mark 305 associated with the diluted concentration of theophylline. Similarly, the therapeutic range for sodium valproate is 50-100 μg / ml, so that a calibration curve can be generated with standard sample concentrations of 20, 50, 100, 200 μg / ml, accompanied by dilution of sodium palproate. A check is given to 20, 50, 100, and 200 μg / ml of the concentration adoption mark 305 that is present.

  The dilution ratio of the dilution series and the setting of the concentration value to be adopted for the calibration curve can be automated. For example, when the analyst instructs the device control unit 207 all the concentration ranges of the components to be measured, among all the components to be measured, the dilution series from the highest concentration to the lowest concentration is a geometric sequence. What is necessary is just to arrange. In this embodiment, since the maximum concentration is 200 μg / ml of sodium palproate and the minimum concentration is 8 μg / ml of theophylline, the concentration range of the component to be measured is included in the logarithmic series based on 1, 2, 5, etc. A dilution series is generated to That is, the series is 5, 10, 20, 50, 100, 200, 500 μg / ml. Next, a series is selected to include each component concentration range. That is, the theophylline may be checked with a concentration adoption mark of 5, 10, 20 μg / ml and sodium parproate of 20, 50, 100, 200, 500 μg / ml. Further, the concentration of the high concentration standard sample may be 500 μg / ml, which is the highest concentration in the series, and all components may be included.

  After the above setting, when the analyst presses the setting button 306, the apparatus control unit 207 stores the setting content and can proceed to the next process (S104). Further, when the number of the standard sample dilution series 303 is not appropriate, the stop button 307 can be pressed to return the process to the previous step.

(Iii) High-concentration standard sample The above-mentioned high-concentration standard sample is prepared as a standard sample containing components having concentrations set to the standard sample pre-dilution concentration 302 in FIG. A high-concentration standard sample is generally prepared by an analyst using a technique. In the present embodiment, all components to be measured may be included in the same concentration most simply. For this reason, it is overwhelmingly simple and has fewer errors than the conventional method in which the concentration of each component is changed in consideration of the measured concentration range. In the present embodiment, as set in FIG. 3, a sample containing theophylline and sodium parproate at 200 μg / ml may be prepared.

(Iv) Standard sample dilution series preparation process (S104)
In the standard sample dilution series preparation process (S104), the apparatus control unit 207 instructs the sample introduction unit 201 to dilute the high concentration standard sample according to the standard sample dilution ratio 304 shown in FIG. This step may be performed by an analyst using a technique, but the sample introduction unit 201 is provided with an automatic dispensing function linked to the apparatus control unit 207 so as to automatically dilute. In addition, you may make it implement | achieve automatic dilution by installing an automatic dispensing apparatus separately from the sample introduction part 201. FIG. When measurement is performed by the internal standard method, an equal amount of the internal standard component is added to all standard samples in this process (S105).

(V) Standard sample measurement process (S106)
In the process from the repetition start of the standard sample number (S105) to the repetition end of the standard sample number (S107), the apparatus control unit 207 produces the number of standard sample dilution series, that is, the standard sample dilution series preparation process (S104). All standard samples are measured (S106). In this step, all the components are measured in all the standard sample dilution series 303 regardless of the application state of the concentration adoption mark 305 shown in FIG. For this reason, in the LC / MS apparatus 200, the sample separation time in the LC apparatus (201 to 202) is dominant in the measurement time, and even if the measurement of only a specific component is omitted, the entire measurement time is not significantly affected. Because.

(Vi) Calibration curve creation process (S108)
In the calibration curve creation process (S108), the apparatus control unit 207 creates a calibration curve using the standard sample measurement results measured in the process from the repetition start of the standard sample number (S105) to the repetition end of the standard sample number (S107). To do.

  FIG. 4 is a schematic diagram of creating a calibration curve according to the first embodiment of the present invention. In each of the two graphs, the concentration when theophylline and sodium parproate are measured is shown on the horizontal axis, and the intensity is shown on the vertical axis. Here, the intensity is, for example, a voltage signal output from an ion detector built in the mass analyzer 204 or A / D converted or a pulse signal count from a light detector. Moreover, when measuring by an internal standard method, it becomes an intensity ratio of an internal standard component and a measurement target component. In this embodiment, an A / D conversion value (ADU) is used as an example. In FIG. 4, both the vertical axis and the horizontal axis are logarithmic graphs.

  The range shown in the background of the graph is the measured dynamic range (DR) 402 of the device. The dynamic range 402 differs depending on the device used and the usage environment. In the measurement of each component, if the signal intensity 401 falls within this range and the measurement is performed with a signal intensity 401 as high as possible, it is less likely to be affected by noise and fluctuations in the background value of the detector. However, if the measurement dynamic range (DR) 402 is exceeded, the signal intensity 401 is saturated and the linearity deteriorates. In the present embodiment, it is assumed that in the analysis condition setting 102, the measurement conditions are optimized so that the measurement dynamic range 402 is used to the maximum with respect to the concentration range where each component is desired to be measured.

  In the measurement results of theophylline, since the measurement conditions are optimized to 8 to 15 μg / ml, which is the concentration range to be measured, the signal intensity 401 is saturated at a concentration of 50 to 200 μg / ml. The theophylline calibration curve 405 is a calibration curve created according to the instructions in FIG. The feature of the present invention is that it is possible to set, for each component, whether or not the measurement intensity is applied to the calibration curve, and the concentration value when it is applied. Thereby, since a calibration curve can be created by selectively using only a standard sample in the vicinity of the concentration range to be measured, the accuracy in the concentration region to be measured is high.

  On each signal intensity 401 in FIG. 4, a circle (◯) mark 403 is displayed for the standard sample to which the concentration adoption mark 305 in FIG. 3 is added, and a cross (×) mark 404 is displayed for the standard sample to which no density is applied. In addition, it is possible to determine at a glance which standard sample the calibration curve is created. A theophylline calibration curve 406 created with all standard samples is a calibration curve created using standard samples of all measured dilution series. It is clearly not accurate because it includes the signal strength in the saturation region. In addition, if the measurement conditions are changed and the component sensitivity is reduced, relatively good linearity can be achieved for all standard samples, but the concentration range to be measured is very close to the origin of the calibration curve. The measurement dynamic range 402 of the apparatus cannot be fully utilized, resulting in a decrease in measurement accuracy.

  Sodium valproate has a therapeutic range of 50 to 100 μg / ml, and the measurement conditions are optimized for this concentration. Therefore, an example is shown in which the signal intensity 401 is below the detection limit at a concentration of 5, 10 μg / ml. Even in this case, if a calibration curve is created in accordance with the instructions in FIG. 3, an accurate sodium valproate calibration curve 407 can be obtained for a desired concentration range. The sodium valproate calibration curve 408 prepared with all the standard samples was prepared including the signal intensity 401 with a concentration of 5 and 10 μg / ml which was below the measurement limit, and is clearly not accurate.

(Vii) Unknown sample measurement process (S110)
In the processing from the repetition start of the unknown sample number (S109) to the repetition end of the unknown sample number (S111), the apparatus control unit 207 measures the unknown sample under the same conditions as the standard sample measurement (S110). The unknown sample in the present embodiment is specifically a pretreatment of the blood of a patient undergoing TDM, and the components set in FIG. 3 are expected to be detected at concentrations within each treatment area. This is a sample.

(Viii) Quantitative calculation processing (S112)
In the quantitative calculation process (S112), the apparatus control unit 207 converts the signal intensity value measured in the unknown sample measurement (S110) into a concentration using the calibration curve created in the calibration curve creation (S108). As shown in FIG. 4, the calibration curve created in the calibration curve creation (S108) is a calibration curve suitable for the concentration region to be measured, and accurate concentration calculation is possible within that range. More specifically, m / z (mass value: horizontal axis) and detection intensity (vertical axis) of each standard sample are obtained by mass spectrometry. Moreover, m / z and detection intensity can be obtained for unknown samples. The detection intensity of the unknown sample is obtained by comparing the graph of the unknown sample and the standard sample. This is because a peak is detected at the same mass for each component. Then, the concentration of each component is obtained using the detected intensity and the obtained calibration curve.

(Ix) Measurement result information output processing (S113)
In the measurement result information output process (S113), the apparatus control unit 207 determines the intensity value of the standard sample measurement (S106), the calibration curve creation result of the calibration curve creation (S108), the intensity value of the unknown sample measurement (S110), and the quantitative calculation. The unknown sample concentration value calculated in (S112) is output to the display device 208. Although detailed explanation is omitted, in the standard sample measurement process (S106) and the unknown sample measurement process (S110), generally, each sample is repeatedly measured to improve accuracy. You may display the average value of this repeated measurement, dispersion | distribution, a relative standard deviation, etc. collectively.

(X) The above processing is performed, and the analysis ends (S114).

<First Embodiment: Summary>
In the first embodiment of the present invention, as shown in the above procedure, a calibration curve can be created by selecting the standard sample intensity near the concentration range to be measured for each component from the standard sample dilution series. An unknown sample can be quantified easily and accurately while reducing the labor and error of production.

(2) Second Embodiment In the first embodiment, theophylline and sodium valproate are exemplified as measurement components, but the difference between the therapeutic ranges of these two drugs is about 10 times at most. For this reason, only six types of dilution series were prepared, and a calibration curve could be created by selecting an appropriate range of concentrations for each component.

  However, for example, tacrolimus has a therapeutic range of 5 to 20 ng / ml, and there is a concentration difference of about 1/10000 of sodium valproate. For this reason, when the method of the first embodiment is used, the range of the dilution series shown in FIG. 3 is increased, and the number of standard samples is increased, so that the entire analysis process takes a long time.

  Therefore, in the second embodiment, the GUI shown in FIG. 3 is improved and a GUI as shown in FIG. 5 is prepared. In the screen (GUI) shown in FIG. 5, the concentration of each component can be designated as the concentration 501 before standard sample dilution. This screen (GUI) is a screen used by the operator (analyzer) for density setting in the measurement density area setting process (S103). Processes other than the measurement density area setting process (S103) in this embodiment are the first. This is the same as the embodiment.

  Theophylline and sodium valproate have the same settings as in the first embodiment, but tacrolimus has an extremely low concentration in the treatment area, so the concentration in the high-concentration standard sample should be kept low before dilution. . It is necessary for the analyst to set which component is contained at what level, depending on the purpose, but it is only necessary that the therapeutic range be included in one of the concentration ranges diluted by the dilution series. The concentration in the high concentration standard sample does not need to be considered so strictly.

  In the example of FIG. 5, 0.1 μg / ml was set as the concentration in the high concentration standard sample. As a result, the concentration of tacrolimus in each dilution series is 0.0025, 0.005, 0.01, 0.025, 0.05, 0.1 μg / ml. In order to quantify the therapeutic range of 5 to 20 ng / ml of tacrolimus, if 0.0025, 0.005, 0.01, 0.025 μg / ml is selected, a calibration curve including the therapeutic range can be created.

(3) Third Embodiment The calibration curve defines its quality with values such as a variation coefficient and a correlation coefficient. The coefficient of variation (CV value) is obtained by repeatedly measuring the components of each standard sample under the same conditions and dividing the standard deviation by the average value. This value is an index indicating how much the measurement results vary. The correlation coefficient is an index indicating how much the strength of each standard sample fits the prepared calibration curve. For example, in fields such as tap water quality analysis, soil environment measurement, and chemical analysis where official analysis methods are established, a coefficient of variation of 10% or less and a correlation coefficient of 0.990 or more are generally required. Further, in fields for research use such as private laboratories and universities, especially in fields where high accuracy is required, a coefficient of variation of 5% or less and a correlation coefficient of about 0.995 are required accuracy indexes.

  In the first and second embodiments, the measurement concentration range setting process (S103) is executed before the standard sample measurement (S106). However, in order to obtain a calibration curve with the required accuracy, it may be necessary to optimize the analysis conditions.

  Therefore, in the third embodiment, after the calibration curve creation process (S108), a process is performed in which the analyst confirms the calibration curve generation state and resets the analysis conditions and the measurement concentration range. FIG. 6 is a flowchart for explaining an analysis procedure according to the third embodiment.

(I) In FIG. 6, the analysis start (S601) to the calibration curve creation process (S608) are equivalent to the processes of S101 to S108 of FIG.

(Ii) In S609, the apparatus control unit 207 determines whether the calibration curve generated in S608 is appropriately created in the treatment area. For example, the device control unit 207 displays the GUI screen shown in FIG. 7 on the display device 208 so that the analyst can check whether the calibration curve of each component is appropriately created with the therapeutic area concentration. FIG. 7 shows an example in which, in addition to the display of FIG. 4, a standard deviation display 708 by error bar display at each standard sample measurement point and a correlation coefficient display 709 at the lower right of the graph are additionally displayed. The larger the error bar 708 spreads, the greater the variance, that is, the greater the variation in the results of repeated measurements. The analyst confirms the degree of duplication of each standard sample to the calibration curve and the numerical values of the standard deviation and correlation coefficient shown in the figure. If the analyst determines that the required accuracy has not been obtained, Standard sample reselection S610 is performed.

  For example, when changing the analysis condition, the analysis condition change button 701 is pressed to display the analysis condition change screen. FIG. 9 is a diagram illustrating an example of an analysis condition change screen. FIG. 9 is an example of a screen for setting measurement conditions in the mass spectrometer 204 of FIG. The setting screen includes an analysis parameter setting area 901 and a mass number calibration area 902 for mass analysis.

  The analysis parameter setting area 901 is an area for examining problems from the analysis results already collected by the analyst and adjusting analysis parameters necessary for improvement.

  The mass number calibration region 902 is a region for calibrating the mass number difference of the mass spectrometer, and by introducing a sample containing a component having a known mass number into the apparatus, the measurement results are sequentially displayed. It has become. The mass number calibration axis 903 can be moved left and right by a coordinate indicator. By moving the mass number calibration axis 903 to the known mass number peak of the sample, an instruction is stored in the apparatus control unit 207 to calibrate the instrumental error, and instrumental error correction is performed.

  In the example of FIG. 7, the first standard sample measurement intensity (intensity at 5 μg / ml) and the second standard sample measurement intensity (intensity at 10 μg / ml) of theophylline are insufficient. It is assumed that the calibration curve 704 is not accurate. Since it is predicted that theophylline component measurement sensitivity is insufficient, it is necessary to correct the measurement conditions so that the appropriate measurement sensitivity is maintained in the treatment area.

  On the other hand, FIG. 9 shows the measurement intensity at each fragment mass number in the mass number calibration region 902 after introducing theophylline and sodium valproate by infusion. In the mass number calibration region 902, for example, the peak position of 124.0, which is the cleavage mass number of theophylline, is shifted from the mass number calibration axis 903 to the low mass number side. For this reason, it is estimated that the sensitivity is lowered. Therefore, the analyst instructs the mass number calibration axis 903 to move to the peak center, changes the analysis condition, presses the remeasurement 904 button, and confirms the change contents of the analysis condition setting. Then, the standard sample measurement process (S606) shown in FIG. 6 is repeated for the number of standard samples under the changed analysis conditions.

(Iii) Next, an example in which only the standard sample reselection is performed in S610 will be described. In the standard sample measurement result of sodium valproate shown in FIG. 7, the signal intensity of the fourth standard sample (intensity when the sodium valproate concentration is 50 μg / ml) is reduced, and error bars, that is, variations in the measurement results are also present. Since it is large, it is considered that the sodium valproate calibration curve 705 is not accurate. If the analyst knows that some sudden problem has occurred during the fourth signal strength measurement, if the standard sample is invalidated, an accurate calibration curve based on the measured signal strength of the other standard sample will be used. Can be created. In this case, the circular (◯) mark 706 drawn on the signal intensity of the fourth standard sample of sodium valproate is indicated with a coordinate indicator or the like, and changed to the cross (×) mark 707, Standard samples can be excluded. Note that, when the standard sample is reselected, it is not necessary to remeasure the sample, so only the calibration curve creation process (S608) needs to be performed.

(Iv) Calibration Curve After Correction FIG. 8 is a diagram showing a calibration curve after correcting the above problem. It can be seen that the theophylline calibration curve 801 and the sodium valproate calibration curve 802 are appropriately prepared in the treatment area of the target component.

  On the other hand, if there is still a problem as a result of the reconfirmation, the analysis condition change / standard sample reselection process (S610) may be repeated until the problem is corrected. When it is confirmed that the problem has been corrected, the measurement continuation button 702 is pressed to proceed to unknown sample measurement. If a problem other than the calibration curve creation occurs and measurement cannot be continued, the cancel button 703 is pressed to stop the measurement.

  In this way, a calibration curve in which the problem is individually corrected for each component can be created while changing the analysis conditions and reselecting the standard sample and confirming the measurement result for each component of the standard sample.

(4) Summary (i) According to the embodiment of the present invention, the analyzer (LC / MS device) includes a plurality of components at arbitrary concentrations (each component may have the same concentration or different concentrations). A GUI (FIG. 3) for instructing the dilution rate of the standard sample for each of the plurality of components is provided on the screen of the display device, and the user sets the dilution rate (plurality) for each component. And an analyzer dilutes a standard sample according to the set dilution rate, and isolate | separates each component. In addition, the analyzer analyzes each component of the concentration corresponding to the instructed dilution rate, measures the intensity of each component of each concentration, and based on this measurement result, generates a calibration curve for each component. Generate. In this way, in the quantitative analysis of a sample containing multiple components, it is not necessary to consider the measurement concentration range of each component when preparing a standard sample, so the effect of reducing the complexity of preparing a standard sample and reducing measurement errors There is.

  The analysis device generates a calibration curve for each component contained in the standard sample based on the intensity information at the concentration corresponding to the instructed dilution rate and the measurement dynamic range of the analysis device. As a result, a calibration curve adapted to the capacity of the analyzer can be generated.

  In addition, the dilution rate prepared in advance in the dilution rate setting GUI displayed on the screen is set to be common to each component. By doing so, the user can set a desired concentration simply by selecting the dilution rate for each component, and thus the load on the user when generating a calibration curve can be reduced. become able to.

  Further, the analyzer includes the generated calibration curve, confirms the suitability of the calibration curve, and changes the analysis conditions when the user determines that the calibration curve is inappropriate. 7) is displayed on the screen of the display device. The analyzer also displays standard deviation information indicating the intensity variation obtained from the past measurement results for the intensity at each concentration of the calibration curve displayed on the calibration curve confirmation GUI. Further, in the calibration curve confirmation GUI, the user can select whether or not to use a combination of each concentration and intensity of each component for creating a calibration curve. When the combination of concentration and intensity used to create the calibration curve is changed by the user, the analyzer again generates a corresponding calibration curve. By doing this, it is possible to confirm the calibration curve once generated and determine that it is inappropriate, so that it can be corrected so as to be appropriate. It is easy.

  In addition, when introducing a sample whose concentration of the contained component is unknown, the analyzer performs mass analysis on the unknown sample, outputs a mass analysis result, and compares this mass analysis result with the generated calibration curve. Thus, the concentration of the unknown sample is specified. Thus, the generated calibration curve is used to specify the concentration of the unknown sample, and since the calibration curve is more accurate, it becomes possible to know a more accurate concentration for the unknown sample.

(Ii) The present invention can also be realized by software program codes that implement the functions of the embodiments. In this case, a storage medium in which the program code is recorded is provided to the system or apparatus, and the computer (or CPU or MPU) of the system or apparatus reads the program code stored in the storage medium. In this case, the program code itself read from the storage medium realizes the functions of the above-described embodiment, and the program code itself and the storage medium storing the program code constitute the present invention. As a storage medium for supplying such program code, for example, a flexible disk, CD-ROM, DVD-ROM, hard disk, optical disk, magneto-optical disk, CD-R, magnetic tape, nonvolatile memory card, ROM Etc. are used.

  Also, based on the instruction of the program code, an OS (operating system) running on the computer performs part or all of the actual processing, and the functions of the above-described embodiments are realized by the processing. May be. Further, after the program code read from the storage medium is written in the memory on the computer, the computer CPU or the like performs part or all of the actual processing based on the instruction of the program code. Thus, the functions of the above-described embodiments may be realized.

  Further, by distributing the program code of the software that realizes the functions of the embodiment via a network, it is stored in a storage means such as a hard disk or memory of a system or apparatus, or a storage medium such as a CD-RW or CD-R And the computer (or CPU or MPU) of the system or apparatus may read and execute the program code stored in the storage means or the storage medium when used.

  Finally, it should be understood that the processes and techniques described herein are not inherently related to any particular apparatus, and can be implemented by any suitable combination of components. In addition, various types of devices for general purpose can be used in accordance with what is described herein. It may prove useful to build a dedicated device to perform the method steps described herein. Various inventions can be formed by appropriately combining a plurality of constituent elements disclosed in the embodiments. For example, some components may be deleted from all the components shown in the embodiment. Furthermore, constituent elements over different embodiments may be appropriately combined. Although the present invention has been described with reference to specific examples, these are in all respects illustrative rather than restrictive. Those skilled in the art will appreciate that there are numerous combinations of hardware, software, and firmware that are suitable for implementing the present invention. For example, the described software can be implemented in a wide range of programs or script languages such as assembler, C / C ++, perl, shell, PHP, Java (registered trademark).

  Furthermore, in the above-described embodiment, control lines and information lines are those that are considered necessary for explanation, and not all control lines and information lines on the product are necessarily shown. All the components may be connected to each other.

  In addition, other implementations of the invention will be apparent to those skilled in the art from consideration of the specification and embodiments of the invention disclosed herein. Various aspects and / or components of the described embodiments can be used singly or in any combination in a computerized storage system capable of managing data. The specification and specific examples are merely exemplary, and the scope and spirit of the invention are indicated in the following claims.

201 ... sample introduction part 202 ... separation part 203 ... ionization part 204 ... mass analysis part 205 ... sample (accommodating part)
206 ... Eluent (container)
207 ... Device control unit 208 ... Display unit 209 ... Operation unit 301 ... Measurement concentration area setting screen 302 ... Standard sample dilution before concentration 303 ... Standard sample dilution series 304 ... Standard Sample dilution ratio 305 ... Concentration adoption mark 306 ... Setting button 307 ... Cancel button

Claims (15)

An analyzer for analyzing a plurality of components contained in a sample,
A standard sample storage unit for storing a standard sample containing each of the plurality of components at a predetermined concentration;
A control unit for displaying a GUI for instructing the dilution rate of the standard sample for each of the plurality of components on the screen of the display device, and receiving the dilution rate of each component instructed by the user;
In response to an instruction from the control unit, a sample introduction unit for introducing the diluted standard sample by diluting the standard sample based on the dilution rate of the respective components;
A separation unit for separating each component from the diluted standard sample, and obtaining a component having a concentration corresponding to the instructed dilution rate;
Analyzing each of the components of the concentration corresponding to the instructed dilution rate, and measuring the intensity of each component of each concentration,
The said control part produces | generates the calibration curve of each component based on the measurement result by the said analysis part, The analyzer characterized by the above-mentioned. In claim 1,
The control unit generates the calibration curve for each component based on the information on the intensity at the concentration corresponding to the instructed dilution rate and the measurement dynamic range of the analysis unit. Analysis equipment. In claim 1,
The dilution ratio prepared in advance in the GUI is set to be common to the respective components,
The standard device includes an analysis device characterized in that each component has an arbitrary concentration. In claim 1,
The control unit further includes a generated calibration curve, confirms the suitability of the calibration curve, and changes the analysis conditions when the user determines that the calibration curve is inappropriate, and a calibration curve confirmation GUI for instructing remeasurement Is displayed on the screen of the display device. In claim 4,
The control unit displays standard deviation information indicating variation in intensity obtained from past measurement results for the intensity at each concentration of the calibration curve displayed on the calibration curve confirmation GUI. . In claim 5,
In the calibration curve confirmation GUI, the user can select whether or not to use a combination of each concentration and intensity of each component for creating a calibration curve,
The said control part produces | generates a corresponding calibration curve again, when the combination of the said density | concentration and intensity | strength used for preparation of the said calibration curve is changed, The analyzer characterized by the above-mentioned. In claim 1,
The sample introduction part introduces a sample whose concentration of the contained component is unknown,
The analysis unit performs mass analysis on the unknown sample and outputs a mass analysis result,
The said control part specifies the density | concentration of the said unknown sample by comparing the said mass spectrometry result and the produced | generated calibration curve, The analyzer characterized by the above-mentioned. A sample analysis method using an analyzer for analyzing a plurality of components contained in a sample,
The analysis apparatus includes a standard sample storage unit that stores a standard sample containing each of the plurality of components at a predetermined concentration, a control unit, a sample introduction unit, a separation unit, and an analysis unit.
The sample analysis method includes:
The control unit displays a GUI for instructing the dilution rate of the standard sample for each of the plurality of components on the screen of the display device, and receiving the dilution rate of each component instructed by the user;
In response to an instruction from the control unit, the sample introduction unit dilutes the standard sample based on the dilution rate of the respective components, and introduces the diluted standard sample;
The separation unit separates each component from the diluted standard sample, and acquires a component having a concentration corresponding to the instructed dilution rate;
The analysis unit analyzing each of the components of the concentration corresponding to the instructed dilution rate, and measuring the intensity of each component of each concentration;
The control unit generates a calibration curve for each component based on the measurement result by the analysis unit;
A sample analysis method characterized by comprising: In claim 8,
In the step of generating the calibration curve, the control unit, for each of the components, based on the intensity information at the concentration corresponding to the instructed dilution rate and the measurement dynamic range of the analysis unit, A sample analysis method characterized by generating a calibration curve. In claim 8,
The dilution ratio prepared in advance in the GUI is set to be common to the respective components,
The sample analysis method, wherein the standard sample contains each component at an arbitrary concentration. In claim 8,
Further, the control unit includes a generated calibration curve, confirms the suitability of the calibration curve, and changes the analysis conditions and instructs remeasurement when the user determines that the calibration curve is inappropriate. A sample analysis method comprising: displaying the image on the screen of the display device. In claim 11,
In the step of displaying the calibration curve confirmation GUI, the control unit, for the intensity at each concentration of the calibration curve displayed on the calibration curve confirmation GUI, shows a standard deviation indicating variation in intensity obtained from past measurement results. A sample analysis method characterized by displaying information. In claim 12,
In the calibration curve confirmation GUI, the user can select whether or not to use a combination of each concentration and intensity of each component for creating a calibration curve,
The sample analysis method further includes the step of generating again a corresponding calibration curve when the control unit changes the combination of the concentration and the intensity used to create the calibration curve. The claim 8, further comprising:
When a sample with an unknown content component concentration is introduced into the sample introduction unit, the analysis unit performs mass analysis on the unknown sample and outputs a mass analysis result; and
The control unit identifying the concentration of the unknown sample by comparing the mass spectrometry result and the generated calibration curve;
A sample analysis method characterized by comprising: A computer-readable storage medium for storing a program for executing a sample analysis method in an analyzer for analyzing a plurality of components contained in a sample,
The analysis apparatus includes a standard sample storage unit that stores a standard sample containing each of the plurality of components at a predetermined concentration, a control unit, a sample introduction unit, a separation unit, and an analysis unit.
The program is
The control unit displays a GUI for instructing the dilution rate of the standard sample for each of the plurality of components on the screen of the display device, and executes a process of receiving the dilution rate of each component instructed by the user Let
Instructing the control unit to cause the sample introduction unit to dilute the standard sample based on the dilution ratio of the respective components and introduce the diluted standard sample,
Causing the control unit to instruct the separation unit to separate each component from the diluted standard sample and obtain a component having a concentration corresponding to the instructed dilution rate;
Instructing the control unit to analyze each of the components of the concentration corresponding to the instructed dilution rate, and to measure the intensity of each component of each concentration,
A computer-readable storage medium characterized by causing the control unit to execute a process of generating a calibration curve of each component based on a measurement result by the analysis unit.

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