JP5962775B2 - Data processing equipment for chromatographic mass spectrometry - Google Patents

Description

  The present invention relates to a data processing apparatus for chromatographic mass spectrometry that processes data collected by a chromatographic mass spectrometer such as a liquid chromatograph mass spectrometer (LC / MS) or a gas chromatograph mass spectrometer (GC / MS). .

  When a sample is analyzed by a liquid chromatograph (LC), a gas chromatograph (GC) or the like, a peak corresponding to a component contained in the sample appears on the chromatogram. Since the size of the peak on the chromatogram depends on the concentration or amount of the component, if the peak corresponds to a single component, the concentration or content of the corresponding component based on the height or area value of the peak. It is possible to determine the quantity. However, the peak that appears to be a single shape on the chromatogram is not necessarily due to a single component, and when multiple components that are not sufficiently separated by the column overlap, Often, a component that does not overlap the target component. Therefore, conventionally, a chromatogram peak purity determination process has been performed to determine whether a peak on a chromatogram is due to a single component.

  For example, Non-Patent Document 1 and Patent Document 1 disclose a peak purity determination processing method in a chromatogram obtained by a liquid chromatograph using a multichannel detector such as a PDA (Photo Diode Array) detector. In this liquid chromatograph, three-dimensional chromatogram data (see FIG. 2A) having three dimensions of time, wavelength, and absorbance is obtained by repeatedly acquiring an absorbance spectrum for the eluate from the column. From the data, an absorbance spectrum at any time point of the peak appearing in the chromatogram can be generated.

Now, since the absorbance spectrum can be regarded as a set of absorbances for each wavelength, certain two absorbance spectra S ₁ and S ₂ are expressed as the following vectors.
↑ S ₁ = (a ₁ (λ ₁ ), a ₁ (λ ₂ ), ..., a ₁ (λ _n ))
↑ S ₂ = (a ₂ (λ ₁ ), a ₂ (λ ₂ ), ..., a ₂ (λ _n ))
Here, a ₁ (λ _i ) and a ₂ (λ _i ) are absorbances at the wavelength λ _i .

また、ベクトル↑Ｓ₁、↑Ｓ₂の各要素で(1)式を表現すると(2)式となる。

(1)式又は(2)式により計算される類似度SIの値が１に近いほど、二つの吸光度スペクトルＳ₁、Ｓ₂の形状はよく一致しているといえる。Assuming that there are only two wavelengths λ ₁ and λ ₂ , two vector-expressed absorbance spectra ↑ S ₁ and ↑ S ₂ start from the origin in a two-dimensional space as shown in FIG. It can be expressed as a vector. When the number of wavelengths is three or more, a vector of the number of wavelengths in a multidimensional space may be considered. If the two absorbance spectra S ₁ and S ₂ are similar in shape, the two vectors ↑ S ₁ and ↑ S ₂ face the same direction, and the angle formed by both vectors ↑ S ₁ and ↑ S ₂ . θ becomes zero. In general, the smaller the angle θ, the higher the degree of similarity of the curve shapes of the two absorbance spectra S ₁ and S ₂ . Therefore, cos θ is calculated and defined as the similarity SI between the two absorbance spectra S ₁ and S ₂ . That is, the similarity SI between two absorbance spectra S ₁ and S ₂ can be obtained from the following equation (1).

Further, when expression (1) is expressed by each element of vectors ↑ S ₁ and ↑ S ₂ , expression (2) is obtained.

It can be said that the shape of the two absorbance spectra S ₁ and S ₂ agrees better as the value of the similarity SI calculated by the equation (1) or (2) is closer to 1.

In the peak purity determination method described in Patent Document 1, for example, the absorbance spectrum at the peak top of the target chromatogram peak that is the target of purity determination is S ₁ , and an arbitrary peak period from the start point to the end point of the target peak is set. the absorbance spectrum obtained by the time and S _2, calculates each similarity SI at each time point. Then, the chromatogram peak area is color-coded according to the similarity SI value so that the change of the calculated similarity SI value with time can be seen at a glance.

  If the target peak is derived only from the target component, the similarity SI is the highest near the peak top and gradually decreases as the distance from the peak top increases, and the shape is generally symmetrical with respect to the central axis of the peak. Become. On the other hand, if there is an impurity in front or behind the peak top of the target peak, one of the front or rear of the peak top of the target peak should have a large decrease in similarity compared to the other. . Thus, the operator can determine that there is a high possibility that impurities are included in the time range in the vicinity thereof.

  However, in the conventional peak purity determination method described above, when a peak of another component having a similar shape of the curve of the absorbance spectrum overlaps with a certain target peak, the degree of decrease in the similarity due to the overlap. Because of the small size, it was difficult to determine whether impurities were included. Further, even if it was found that the impurity overlapped the target peak, it was impossible to estimate what the impurity was.

  In particular, when performing quantitative analysis of a component in a sample containing a single component, a flow injection analysis (FIA = Flow Injection Analysis) method that does not use a column (that is, does not perform component separation) should be used. There is. The FIA method is a method in which a predetermined amount of sample is injected into a mobile phase fed at a constant flow rate using a liquid chromatograph injector or the like, and the sample is introduced into a detector by being placed on the flow of the mobile phase. Yes, as with the eluate from the column outlet, the concentration of the target component changes in an approximately mountain shape over time. The data obtained when a sample introduced by the FIA method is detected by a multichannel detector is also three-dimensional data having three dimensions of time, wavelength, and absorbance, and is a liquid chromatograph as described above. Is substantially the same as the data collected by Therefore, the “chromatograph mass spectrometer” as used in this specification includes not only a chromatograph column but also an apparatus for introducing a sample into the mass spectrometer by the FIA method.

Japanese Patent No. 2936700

Yasutaka Mito, Mitsuo Kitaoka, "Photodiode Array UV-VIS Detector SPD-M6A for Shimadzu HPLC", Shimazu Review, Vol. 46, No. 1, July 1989, pp.21-28

  The present invention has been made in order to solve the above-mentioned problems. The main purpose of the present invention is that the peak of another component having a shape similar to that of the target compound in the absorbance spectrum is present relative to the chromatogram peak of the target compound. It is an object of the present invention to provide a data processing apparatus for chromatographic mass spectrometry capable of performing a highly reliable peak purity determination, which can identify the overlap of impurities even when they overlap.

  Another object of the present invention is to provide a data processing apparatus for chromatographic mass spectrometry that can estimate what a component is when it is found that another component overlaps the chromatogram peak of the target compound. Is to provide.

In order to achieve the above object, the first aspect of the present invention provides data for processing data collected by a chromatograph mass spectrometer combining a chromatograph having a multichannel detector and a mass spectrometer. A processing device comprising:
a) First to detect a peak derived from a target compound in a chromatogram created based on the first three-dimensional chromatogram data collected by the multi-channel detector and having dimensions of time, wavelength, and absorbance. A peak detector;
b) Detecting a peak derived from the target compound in a chromatogram created based on the second three-dimensional chromatogram data collected by the mass spectrometer and having dimensions of time, mass-to-charge ratio, and signal intensity. A second peak detector;
c) Regarding the peak derived from the target compound detected by the first peak detector, the absorbance spectrum to be evaluated at each other time point during the period in which the peak appears, and the reference absorbance spectrum at the peak top of the peak or the evaluation object A first similarity calculator that calculates the similarity of the pattern with the reference absorbance spectrum at a time point away from the absorbance spectrum for a predetermined period;
d) For the peak derived from the target compound detected by the second peak detector, the evaluation target mass spectrum at each point in time during which the peak appears, and the reference mass spectrum at the peak top of the peak or the evaluation target mass A second similarity calculator for calculating the similarity of the pattern with the reference mass spectrum at a time point away from the spectrum for a predetermined period;
e) A graph in which the similarity at each time point calculated by the first similarity calculation unit is plotted on the time axis, and a similarity in each time point calculated by the second similarity calculation unit is plotted on the time axis. A similarity evaluation information presentation unit that creates a graph related to the two similarities and displays the graphs on the display screen in a comparable manner;
It is characterized by having.

The second aspect of the present invention, which has been made to achieve the above object, processes data collected by a chromatograph mass spectrometer that is a combination of a chromatograph having a multi-channel detector and a mass spectrometer. A data processing device,
a) First to detect a peak derived from a target compound in a chromatogram created based on the first three-dimensional chromatogram data collected by the multi-channel detector and having dimensions of time, wavelength, and absorbance. A peak detector;
b) Detecting a peak derived from the target compound in a chromatogram created based on the second three-dimensional chromatogram data collected by the mass spectrometer and having dimensions of time, mass-to-charge ratio, and signal intensity. A second peak detector;
c) Regarding the peak derived from the target compound detected by the first peak detector, the absorbance spectrum to be evaluated at each other time point during the period in which the peak appears, and the reference absorbance spectrum at the peak top of the peak or the evaluation object A first similarity calculator that calculates the similarity of the pattern with the reference absorbance spectrum at a time point away from the absorbance spectrum for a predetermined period;
d) For the peak derived from the target compound detected by the second peak detector, the evaluation target mass spectrum at each point in time during which the peak appears, and the reference mass spectrum at the peak top of the peak or the evaluation target mass A second similarity calculator for calculating the similarity of the pattern with the reference mass spectrum at a time point away from the spectrum for a predetermined period;
e) A graph in which the similarity at each time point calculated by the first similarity calculation unit is plotted on the time axis, and a similarity in each time point calculated by the second similarity calculation unit is plotted on the time axis. A determination unit for determining whether or not the peak derived from the target compound is derived from a single component, based on
It is characterized by having.

  Here, the “chromatograph” may be either a liquid chromatograph or a gas chromatograph. In addition, “multi-channel detectors” generally have a combination of a spectroscope and a photodiode array (PDA) detector that detects wavelength-dispersed light at the same time. As long as it is broad (slow change), UV-visible spectrophotometer, infrared spectrophotometer, near-infrared spectrophotometer, fluorescence spectrophotometer with wavelength scanning to obtain absorbance spectrum , Etc.

  In addition, the “chromatograph mass spectrometer” separates components by the FIA method using a liquid chromatograph injector or the like instead of introducing a sample through a column for separating components in a chromatograph into the mass spectrometer. A sample that is not present may be introduced into the mass spectrometer.

  In addition, the “chromatogram created based on the first three-dimensional chromatogram data” is usually a mass chromatogram (also referred to as “extracted ion chromatogram”) at a specific mass-to-charge ratio. A total ion chromatogram may be used.

  In the data processing apparatus for chromatographic mass spectrometry according to the first and second aspects of the present invention, the similarity of the absorbance spectrum pattern conventionally used for determining the purity of the peak derived from the target compound appearing in the chromatogram. The similarity of the pattern of the mass spectrum obtained by the mass spectrometer is used in addition to the property.

  That is, when SIM (Selection Ion Monitoring) measurement for a plurality of mass-to-charge ratios or scan measurement for a predetermined mass-to-charge ratio range is executed in a mass spectrometer, three-dimensional chromatogram data having dimensions of time, mass-to-charge ratio, and signal intensity. Therefore, the second peak detector generates a mass chromatogram based on the three-dimensional chromatogram data, for example, in the mass-to-charge ratio corresponding to the target compound (typically, the quantitative ion characterizing the target compound), A peak near the retention time of the target compound is detected on the chromatogram. For the detected peak derived from the target compound, the second similarity calculation unit calculates a mass spectrum to be evaluated at a certain point in time during which the peak appears, and a reference at a point apart from the mass spectrum to be evaluated for a predetermined period. The process of calculating the similarity of the pattern with the mass spectrum (or the reference mass spectrum at the peak top) is repeatedly executed for all the time points in the peak period or at least a part of the time points.

  Note that when a mass spectrum at a time point away from the evaluation target mass spectrum for a predetermined period is used as the reference mass spectrum, it is preferable that the operator can appropriately set the value of the predetermined period. This is because it is preferable that the predetermined period can be adjusted according to the temporal spread of the impurities overlapping the peak derived from the target compound.

  In the chromatograph mass spectrometry data processing device according to the first aspect of the present invention, the similarity evaluation information presentation unit plots the similarity at each time point calculated based on the absorbance spectrum on the time axis, Create a graph plotting the similarity at each time point calculated based on the mass spectrum on the time axis, and compare the graphs of these two similarities on the display screen, for example, overlaying on the same time axis indicate.

  Even when the target compound and the impurity have similar absorbance spectrum patterns, it is rare that the target compound and the impurity have similar mass spectrum patterns. Conversely, even if the target compound and the impurity have similar mass spectrum patterns, it is rare that the target compound and the impurity have similar absorbance spectrum patterns. Therefore, when an impurity overlaps the peak derived from the target compound, there is a high possibility that a drop in similarity is observed in either of the two graphs related to similarity. Therefore, the operator can accurately and intuitively determine whether or not there is an overlap of impurities in the peak derived from the target compound by visually confirming the displayed graphs regarding two different degrees of similarity.

  On the other hand, in the chromatograph mass spectrometry data processing apparatus according to the second aspect of the present invention, the determination unit executes determination processing in place of the operator's determination as described above. Specifically, for example, when the slope of the curve indicating the temporal change in similarity or the rate of change is calculated, and there is a drop in which the degree of similarity reduction based on such calculation results is greater than or equal to a threshold value, It is determined that there is an overlap. Of course, a determination unit that performs such automatic determination and a similarity evaluation information presentation unit that displays a graph relating to two similarities may be provided.

In the chromatograph mass spectrometry data processing apparatus according to the first and second aspects of the present invention, preferably,
On the graph based on the similarity by the second similarity calculation unit, the mass spectrum near the peak top of the negative peak where the similarity falls, and before the fall of the peak near the negative peak or after the rise A subtraction processing unit for obtaining a difference mass spectrum from the mass spectrum at the time point;
An impurity estimator for estimating a component corresponding to the negative peak based on the difference mass spectrum;
It is good to set it as the structure further provided.
In addition, since the peak intensity on the mass spectrum depends on the concentration (amount) of the contained component at each time point, when calculating the differential mass spectrum, the difference in the concentration of the target compound in the two mass spectra may be corrected.

  Ideally, the difference mass spectrum is an impurity-derived mass spectrum, so the impurity estimation unit qualifies the impurity by, for example, checking the pattern of the difference mass spectrum with the mass spectrum pattern in the compound qualitative database. . Of course, since the approximate retention time of the impurity is also found from the graph relating to the similarity, the information on the retention time can also be used when qualifying the impurity. This not only reveals that the impurities are overlapping, but also makes it possible to know the components.

Since noise peaks due to various factors appear in the mass spectrum, in the chromatograph mass spectrometry data processing apparatus according to the first and second aspects of the present invention, the second similarity calculation unit calculates the similarity. It is preferable to execute a process of removing background noise of each mass spectrum before.

In general, in an ion source of a mass spectrometer, ions having various mass-to-charge ratios are generated from one compound, and if only one or a few characteristic ions are captured, the presence or absence of the compound can be determined. It is. Therefore, in the data processing apparatus for chromatograph mass spectrometry according to the first and second aspects of the present invention, the second similarity calculation unit calculates a peak below a predetermined threshold for each mass spectrum before calculating the similarity. It is advisable to execute the removal process.

  By removing background noise and other unnecessary peaks as described above prior to similarity calculation, the mass spectrum pattern is simplified while retaining the characteristics of the contained compounds. The calculation process is simplified and the time required for this can be reduced. Moreover, the accuracy of similarity calculation itself can be improved, and the accuracy of peak purity determination can be improved.

According to the data processing apparatus for chromatographic mass spectrometry according to the present invention, it is possible to determine the purity of the peak appearing in the chromatogram with higher reliability than before. In particular, since impurities that cannot be detected by conventional peak purity determination methods can be detected, the accuracy of peak purity determination is greatly improved.
Further, according to the preferred configuration in which the subtraction processing unit and the impurity estimation unit are added in the chromatograph mass spectrometry data processing device according to the present invention, it is possible to simply determine whether or not the impurity is overlapped with the peak derived from the target compound. In addition to being able to do so, it becomes possible to know what the impurities are.

The block diagram of the principal part of the liquid chromatograph mass spectrometer (LC / MS) provided with the data processor for chromatograph mass spectrometry which is one Example of this invention. It is a conceptual diagram which shows an example of the three-dimensional chromatogram data obtained by LC / MS of a present Example, (a) is the data obtained with a PDA detector, (b) is the data obtained with a mass spectrometer. The figure which shows the mass chromatogram for the peak purity determination method in LC / MS of a present Example, and the mass spectrum in the specific time. Explanatory drawing of the mass spectrum similarity calculation method at the time of performing peak purity determination in LC / MS of a present Example. The figure which shows the example of a display of the similarity graph for peak purity determination in LC / MS of a present Example. Explanatory drawing of the similarity of two absorbance spectra for chromatogram peak purity judgment.

Hereinafter, an example of an LC / MS provided with a data processing apparatus for chromatographic mass spectrometry according to the present invention will be described in detail with reference to the accompanying drawings.
FIG. 1 is a configuration diagram of a main part of the LC / MS of this embodiment, and FIG. 2 is a conceptual diagram showing an example of three-dimensional chromatogram data obtained by the LC / MS of this embodiment.

  The LC unit 1 includes a mobile phase container 11, a liquid feed pump 12, a sample injection unit 13, a column 14, and a PDA detector 15. The liquid feed pump 12 sucks the mobile phase from the mobile phase container 11 and sends it to the sample injection unit 13 at a constant flow rate. The sample injection unit 13 injects a sample into the mobile phase at a predetermined timing. The injected sample rides on the flow of the mobile phase and is sent to the column 14. While passing through the column 14, each component in the sample is separated and eluted in the time direction. The PDA detector 15, which is a kind of multi-channel detector, irradiates the eluent with light from a light source (not shown), wavelength-disperses the light transmitted through the eluate, and determines the intensity of light at each wavelength by the PDA linear sensor. Detect almost simultaneously. The detection signal repeatedly obtained by the PDA detector 15 is converted into a digital signal by the A / D converter 17 and then three-dimensional chromatogram data having three dimensions of time, wavelength, and absorbance (see FIG. 2A). Is output to the data processing unit 2.

  The eluate that has passed through the PDA detector 15 (or a part of the eluate branched before the PDA detector 15) is introduced into an atmospheric pressure ionization mass spectrometer 16 provided at the rear stage of the LC unit 1. The atmospheric pressure ionization mass spectrometer 16 includes a mass analyzer such as an ion source or a quadrupole mass filter by, for example, an electrospray ionization method or an atmospheric pressure chemical ionization method, and ionizes and generates components in the introduced eluate. The detected ions are separated and detected at a mass-to-charge ratio m / z. The detection signal repeatedly obtained by the atmospheric pressure ionization mass spectrometer 16 is converted into a digital signal by the A / D converter 18 and then three-dimensional chromatogram data having three dimensions of time, mass-to-charge ratio, and signal intensity ( The data is output to the data processing unit 2 as shown in FIG.

  The data processing unit 2 outputs the UV data storage unit 20 for storing the three-dimensional chromatogram data (hereinafter referred to as “UV data”) output from the A / D converter 17 and the A / D converter 18. MS data storage unit 21 for storing three-dimensional chromatogram data (hereinafter referred to as “MS data”), a wavelength chromatogram representing a time change of absorbance at a predetermined wavelength, or a signal intensity time at a predetermined mass-to-charge ratio A chromatogram creation unit 22 that creates a mass chromatogram representing a change, a peak detection unit 23 that detects a peak corresponding to the target compound indicated by the operator in the wavelength chromatogram or the mass chromatogram, and a target compound Measure the absorbance spectrum or mass spectrum similarity at each point in time during the peak period. The similarity calculation unit 24, the UV similarity storage unit 25 that stores the similarity of the absorbance spectrum, the MS similarity storage unit 26 that stores the similarity of the mass spectrum, and the time of the similarity between the absorbance spectrum and the mass spectrum A degree-of-similarity graph creation unit 27 for creating a graph showing a change in the degree of change; a peak purity determination unit 28 for judging whether or not an impurity exists in the peak derived from the target compound based on the created degree-of-similarity graph; An impurity identification unit 29 that identifies an impurity overlapping a peak derived from the target compound based on the graph is included as a functional block.

In addition, the analysis control unit 19 controls the operation of each unit of the LC unit 1, the atmospheric pressure ionization mass spectrometer 16, and the like in order to perform LC / MS analysis on the sample. The control unit 3 controls the entire system and user interface higher than the analysis control unit 19, and is connected to an operation unit 4 operated by an operator and a display unit 5 such as a monitor.
Note that some or all of the functions of the data processing unit 2, the control unit 3, and the analysis control unit 19 are achieved by executing dedicated control / processing software installed in a personal computer or workstation.

  When the target compound is quantitatively analyzed by the LC / MS of this example, one or more mass-to-charge ratios characterizing the target compound are instructed by the operator, and the atmospheric pressure ionization mass spectrometer 16 uses the mass-to-charge ratio. SIM measurement and scan measurement over a predetermined mass-to-charge ratio range are repeated at least within a predetermined time range near the retention time during which the compound elutes. Since mass spectrum data over a predetermined mass-to-charge ratio range is obtained for each scan measurement, as shown in FIG. 2 (b), the MS data storage unit 21 has one set for each predetermined time interval Δt2. Mass spectrum data is stored. On the other hand, since the PDA detector 15 obtains absorbance spectrum data over a predetermined wavelength range for each measurement (one cycle), the UV data storage unit 20 stores a predetermined spectrum as shown in FIG. One set of absorption spectrum data is stored for each time interval Δt1. Normally, Δt1 and Δt2 are not the same.

As shown in FIGS. 2 (a) and 2 (b), the chromatogram derived from the target compound is executed mainly by the data processing unit 2 in a state where the UV data and the MS data are stored in the storage units 20 and 21, respectively. The peak purity determination process will be described in detail with reference to FIGS.
FIG. 3 is a diagram illustrating an example of a mass chromatogram for explaining a peak purity determination method and a mass spectrum at a specific time point, FIG. 4 is an explanatory diagram of a method for calculating a mass spectrum similarity, and FIG. 5 is a similarity for peak purity determination. It is a figure which shows the example of a display of a degree graph.

  For example, when an operator designates a target compound from the operation unit 4, the chromatogram creation unit 22 receives this and reads MS data in a predetermined time range near the retention time of the designated target compound from the MS data storage unit 21. The signal intensity at each time point in the mass-to-charge ratio of ions (generally called “quantitative ions”) that characterize the target compound most is extracted, and a mass chromatogram for the mass-to-charge ratio is created. Next, the peak detector 23 detects the peak near the holding time by examining the change in signal intensity in the time direction in the mass chromatogram, and obtains the time of the peak start point, peak end point, peak top, and the like. For example, the slope amount of the curve of the mass chromatogram shown on the right side of FIG. 2 (b) is sequentially examined in the time direction, and when the slope amount becomes a predetermined value or more, it is determined that it is the peak start point. Is determined to be the peak top when the value changes from positive to zero and further changes to the negative value, and when the absolute value of the inclination amount becomes a predetermined value or less, it is determined that the peak end point is reached.

  The mass chromatogram of quantitative ions is not created from data collected by scan measurement, but from data collected by SIM measurement targeting quantitative ions performed in parallel with scan measurement. May be. In general, when calculating a peak area value for quantification, a mass chromatogram of quantitative ions obtained by SIM measurement is used, but the chromatogram used for peak purity determination is not limited thereto.

Next, the similarity calculation unit 24 calculates the similarity of the mass spectrum obtained at each time point during the peak period from the start point to the end point of the detected peak as follows.
As an example, it is assumed that a mass chromatogram at m / z 307 derived from the target compound is obtained as shown in FIG. 3, and that peak detection is performed for this and the peak period P is determined. FIG. 3 also shows a mass spectrum at each time of the peak start point, peak end point, and peak top included in this peak period, and a mass spectrum at a certain time outside the peak period (baseline mass spectrum). Yes.

Since the horizontal axis of the mass spectrum, that is, the range of the mass-to-charge ratio, is determined at the time of scanning measurement, the mass spectrum is also a signal for each mass-to-charge ratio, just as the absorbance spectrum can be regarded as a collection of absorbances for each wavelength. It can be regarded as a set of strengths. Therefore, the two mass spectra Q ₁ and Q ₂ are expressed as vectors composed of the following elements.
↑ Q ₁ = (b ₁ (m / z ₁ ), b ₁ (m / z ₂ ), ..., b ₁ (m / z _n ))
↑ Q ₂ = (b ₂ (m / z ₁ ), b ₂ (m / z ₂ ), ..., b ₂ (m / z _n ))
Here, b ₁ (m / z _i ) and b ₂ (m / z _i ) are signal intensity values at the mass to charge ratio m / z _i .

  However, the change in signal intensity in the horizontal axis direction (mass-to-charge ratio direction in the mass spectrum and wavelength direction in the absorbance spectrum) is sharper in the mass spectrum than in the absorbance spectrum. In other words, the mass spectrum is in comparison with the absorbance spectrum. Although the resolution is high, noise peaks also appear relatively clearly. Therefore, in calculating the similarity based on the mass spectrum, it is desirable to remove the background noise data of the mass spectrum in advance. This is done by using a background noise removal process that is generally performed on a mass spectrum, for example, a process of subtracting a mass spectrum obtained when no component is present as a baseline mass spectrum. That's fine.

  Also, usually, a plurality of ions having different mass-to-charge ratios are generated from one compound, and depending on the case (for example, depending on ionization conditions, etc.), there are many types, and the calculation of similarity becomes complicated. Therefore, it is preferable to set an appropriate threshold value for the signal intensity in advance, and to exclude data whose signal intensity is equal to or lower than the threshold value from the calculation by setting the intensity value to 0. In addition to the removal of the background noise, by performing such an unnecessary peak data exclusion process, the number of elements constituting the vector is reduced, the similarity can be easily calculated, and the calculation speed can be increased. it can.

このマススペクトルの類似度SI(MS)の値が１に近いほど、二つのマススペクトルＱ₁、Ｑ₂のパターンはよく一致しているといえる。Similar to the similarity of the absorbance spectrum, the more similar the patterns of the two mass spectra Q ₁ and Q ₂ , the closer to zero the angle θ between the two vectors ↑ Q ₁ and ↑ Q ₂ . Therefore, here, the similarity SI (MS) between two mass spectra is defined by the following equation (3) similar to equation (2).

The closer the value of the mass spectrum similarity SI (MS) is to 1, the more closely the patterns of the two mass spectra Q ₁ and Q ₂ match.

  As described above, here, the similarity SI (MS) of the mass spectrum at each time point during the peak period is obtained. Therefore, as shown in FIG. 4, the similarity of the mass spectrum M1 to be evaluated obtained at a certain time t1 during the peak period goes back by n scans (that is, n mass spectra) from that time. The mass spectrum M2 obtained at time t2 is calculated as the reference mass spectrum. That is, first, the mass spectrum at the start point of the peak is set as the mass spectrum M1 to be evaluated, and the baseline mass spectrum obtained at a time point that is back by n scans is set as the reference mass spectrum M2. The similarity SI (MS) is calculated by the equation (3).

Next, the mass spectrum after one scan from the peak start point is set as the evaluation target mass spectrum M1, and the reference mass spectrum is also shifted backward by one scan (that is, the value of n is maintained). Calculate similarity SI (MS). This is repeated until the reference mass spectrum reaches the peak end point, and the similarity SI (MS) of the mass spectrum included in the entire range during the peak period is calculated. In an ideal state where there is no overlap of impurities in the peak derived from the target compound, the similarity should be almost 1 when the two mass spectra Q ₁ and Q ₂ are both in the peak period. If there is, the similarity decreases in a time range in which the overlap exists.

In the example shown in FIG. 3, if the signal intensity threshold for removing unwanted peaks is set to 25,000, for example, only two peaks of m / z 238 and m / z 420 remain in the baseline mass spectrum, and other peaks remain. Are all deleted. That is, the vector representation of the baseline mass spectrum is ↑ Q ₁ = (b ₁ (m / z 238), b ₁ (m / z 420), for example, the background for the mass spectrum at the peak start point or peak top. When the above-mentioned baseline mass spectrum is subtracted as noise removal processing, only the peak at m / z 307 characterizing the target compound remains as the peak with high signal intensity, so there is a large signal intensity peak derived from impurities. Otherwise, when both mass spectra enter the peak period, only the peak at m / z 307 should appear in the mass spectra, and the similarity SI (MS) is almost 1. On the other hand, m / z 307 If there is a peak at a mass-to-charge ratio other than, the similarity SI (MS) will be lowered. Similarity at each time point corresponding to the peak of the compound SI (MS) is temporarily stored in the MS similarity storage unit 26.

  The value of “n” that determines the time interval between the two mass spectra when calculating the similarity is preferably determined according to the width of the impurity peak. Therefore, it is preferable that the operator can make an input setting from the operation unit 4.

  Next, the chromatogram creation unit 22 reads out UV data in a predetermined time range near the retention time of the designated target compound from the UV data storage unit 20, and calculates, for example, the absorbance at each time point at a typical absorption wavelength of the target compound. Extract and create a wavelength chromatogram. Next, the peak detection unit 23 detects the peak near the retention time in the wavelength chromatogram in the same manner as performed for the mass chromatogram, and obtains the time of the peak start point, peak end point, peak top, and the like. .

The similarity calculation unit 24 calculates the similarity of the absorbance spectrum obtained at each time point during the peak period from the start point to the end point of the detected peak, similarly to the similarity to the mass spectrum. That is, as described above, the two absorbance spectra S ₁ and S ₂ are expressed as vectors, and the similarity SI (UV) between the two absorbance spectra S ₁ and S ₂ is defined by the above equation (2). Similarly to the mass spectrum similarity SI (MS), the absorbance spectrum similarity SI (UV) at each time point during the peak period on the wavelength chromatogram is calculated. The similarity SI (UV) also approaches 1 as the two absorbance spectrum curves are closer. Thus, the similarity SI (UV) at each time point corresponding to the peak of the target compound calculated based on the UV data is temporarily stored in the UV similarity storage unit 25. That is, for the same target compound, the mass spectrum similarity SI (MS) and the absorbance spectrum similarity SI (UV) in the vicinity of the appearance period of the chromatogram peak are obtained.

  The similarity graph creation unit 27 reads the similarity data stored in the UV similarity storage unit 25 and the similarity data stored in the MS similarity storage unit 26, respectively, and the horizontal axis represents time and the vertical axis Create a graph with the similarity. Then, the two similarity graphs are superimposed on the same graph frame and drawn in different display colors, for example, and displayed on the screen of the display unit 5 through the control unit 3 (see FIG. 5). Even if the composition or molecular structure of the impurity overlapping the peak derived from the target compound is completely different from that of the target compound, the light absorption characteristics of the impurity may be similar to the light absorption characteristics of the target compound. In such a case, even if the similarity SI (UV) of the absorbance spectrum is close to 1, the similarity SI (MS) of the mass spectrum reflecting the difference in composition and structure is lower than 1. Conversely, even if the impurity composition and structure are similar to the target compound, the light absorption characteristics may be different. In this case, the similarity SI (MS) of the mass spectrum is close to 1, but the similarity SI (UV) of the absorbance spectrum reflecting the difference in the absorption characteristics is lower than 1.

  In the display example of FIG. 5, a clear drop, that is, a negative peak can be observed in the similarity SI (MS) of the mass spectrum as shown in a region C surrounded by a dotted line. During the period in which this negative peak occurs, although there is no drop in the similarity SI (UV) of the absorbance spectrum, the overlap of impurities is suspected. The operator can visually check such a similarity graph and judge the presence or absence of impurities by, for example, empirically judging the degree of depression.

  Further, the peak purity determination unit 28 automatically determines the purity of the peak based on the temporal change amount of the similarity in the two similarity graphs. If there is an overlap of impurities in the peak derived from the target compound, a negative peak appears in any similarity SI (MS) or SI (UV) as shown in FIG. What is necessary is just to determine the presence or absence of impurities by calculating the difference between the similarity and the similarity immediately before the start point of the negative peak or immediately after the end point, and comparing this difference (or rate of change, etc.) with a threshold value. . Displaying such an automatic peak purity determination result on the display unit 5 can assist the operator's determination. Of course, automatic peak purity determination may be performed without displaying the similarity graph, and only the result may be displayed.However, by displaying the similarity graph together, the operator can visually determine whether the automatic determination is appropriate. I can confirm.

When the peak purity determination unit 28 determines that impurities are mixed, or when the operator determines that impurities are mixed as a result of observing the similarity graph, and instructs the fact from the operation unit 4 as follows: Thus, the impurity identification unit 29 executes component identification of the impurity.
Since impurity identification is possible only when a negative peak is observed in the similarity graph of the mass spectrum, it is concluded that impurity identification is not possible unless this negative peak is observed. If a negative peak is observed in the similarity graph of the mass spectrum, the mass at an appropriate point in time when the similarity with the mass spectrum at the peak top of the negative peak is close to 1 and out of the negative peak period. Acquire the spectrum. Then, after multiplying one mass spectrum by a predetermined magnification so as to align the intensity of the characteristic peak derived from the target compound (usually the peak of the maximum intensity appearing in the mass spectrum when the similarity is close to 1), The difference mass spectrum which took the difference of both is calculated.

  In this differential mass spectrum, the peak derived from the target compound disappears and the peak derived from the impurity remains. That is, since the differential mass spectrum is equivalent to the mass spectrum for the impurity, the impurity identifying unit 29 identifies the impurity by comparing the spectrum pattern of the differential mass spectrum with a component qualitative database provided in advance. In this identification, the approximate retention time of impurities estimated from the appearance position of the negative peak in the similarity graph can also be used. In addition, when definitive identification cannot be performed, a plurality of components may be extracted as candidates. Such impurity identification results are also displayed on the screen of the display unit 5 through the control unit 3. Thereby, when the impurity overlaps the peak derived from the target compound, it is possible to inform the operator what the impurity is.

  It should be noted that the above embodiment is an example of the present invention, and it is obvious that modifications, additions, and modifications as appropriate within the scope of the present invention are included in the scope of the claims of the present application.

  For example, although the said Example is an example which applied this invention to LC / MS, it is also applicable to GC / MS whose chromatograph is a gas chromatograph. However, a chromatograph using a multi-channel detector such as a PDA detector is usually a liquid chromatograph. In addition, not the data obtained by detecting the sample separated by the column of the chromatograph with the detector, but the components in the sample introduced without being separated by the FIA method, the multichannel detector and the mass spectrometer. As described above, the present invention can also be applied to an apparatus for processing data obtained by detection in (1).

DESCRIPTION OF SYMBOLS 1 ... Liquid chromatograph (LC) part 11 ... Mobile phase container 12 ... Liquid feed pump 13 ... Sample injection part 14 ... Column 15 ... PDA detector 16 ... Atmospheric pressure ionization mass spectrometer 17, 18 ... A / D converter 19 Analytical control unit 2 Data processing unit 20 UV data storage unit 21 MS data storage unit 22 Chromatogram creation unit 23 Peak detection unit 24 Similarity calculation unit 25 UV similarity storage unit 26 MS similarity Storage unit 27 ... Similarity graph creation unit 28 ... Peak purity determination unit 29 ... Impurity identification unit 3 ... Control unit 4 ... Operation unit 5 ... Display unit

Claims (5)

A data processing device for processing data collected by a chromatograph mass spectrometer combining a chromatograph having a multichannel detector and a mass spectrometer,
a) First to detect a peak derived from a target compound in a chromatogram created based on the first three-dimensional chromatogram data collected by the multi-channel detector and having dimensions of time, wavelength, and absorbance. A peak detector;
b) Detecting a peak derived from the target compound in a chromatogram created based on the second three-dimensional chromatogram data collected by the mass spectrometer and having dimensions of time, mass-to-charge ratio, and signal intensity. A second peak detector;
c) Regarding the peak derived from the target compound detected by the first peak detector, the absorbance spectrum to be evaluated at each other time point during the period in which the peak appears, and the reference absorbance spectrum at the peak top of the peak or the evaluation object A first similarity calculator that calculates the similarity of the pattern with the reference absorbance spectrum at a time point away from the absorbance spectrum for a predetermined period;
d) For the peak derived from the target compound detected by the second peak detector, the evaluation target mass spectrum at each point in time during which the peak appears, and the reference mass spectrum at the peak top of the peak or the evaluation target mass A second similarity calculator for calculating the similarity of the pattern with the reference mass spectrum at a time point away from the spectrum for a predetermined period;
e) A graph in which the similarity at each time point calculated by the first similarity calculation unit is plotted on the time axis, and a similarity in each time point calculated by the second similarity calculation unit is plotted on the time axis. A similarity evaluation information presentation unit that creates a graph related to the two similarities and displays the graphs on the display screen in a comparable manner;
A data processing apparatus for chromatographic mass spectrometry, comprising: A data processing device for processing data collected by a chromatograph mass spectrometer combining a chromatograph having a multichannel detector and a mass spectrometer,
a) First to detect a peak derived from a target compound in a chromatogram created based on the first three-dimensional chromatogram data collected by the multi-channel detector and having dimensions of time, wavelength, and absorbance. A peak detector;
b) Detecting a peak derived from the target compound in a chromatogram created based on the second three-dimensional chromatogram data collected by the mass spectrometer and having dimensions of time, mass-to-charge ratio, and signal intensity. A second peak detector;
c) Regarding the peak derived from the target compound detected by the first peak detector, the absorbance spectrum to be evaluated at each other time point during the period in which the peak appears, and the reference absorbance spectrum at the peak top of the peak or the evaluation object A first similarity calculator that calculates the similarity of the pattern with the reference absorbance spectrum at a time point away from the absorbance spectrum for a predetermined period;
d) For the peak derived from the target compound detected by the second peak detector, the evaluation target mass spectrum at each point in time during which the peak appears, and the reference mass spectrum at the peak top of the peak or the evaluation target mass A second similarity calculator for calculating the similarity of the pattern with the reference mass spectrum at a time point away from the spectrum for a predetermined period;
e) A graph in which the similarity at each time point calculated by the first similarity calculation unit is plotted on the time axis, and a similarity in each time point calculated by the second similarity calculation unit is plotted on the time axis. A determination unit for determining whether or not the peak derived from the target compound is derived from a single component, based on
A data processing apparatus for chromatographic mass spectrometry, comprising: A data processing apparatus for chromatographic mass spectrometry according to claim 1 or 2,
On the graph based on the similarity by the second similarity calculation unit, the mass spectrum near the peak top of the negative peak where the similarity falls, and before the fall of the peak near the negative peak or after the rise A subtraction processing unit for obtaining a difference mass spectrum from the mass spectrum at the time point;
An impurity estimator for estimating a component corresponding to the negative peak based on the difference mass spectrum;
A data processing apparatus for chromatographic mass spectrometry, further comprising:
A data processing apparatus for chromatographic mass spectrometry according to any one of claims 1 to 3,
The data processing apparatus for chromatographic mass analysis, wherein the second similarity calculation unit executes a process of removing background noise of each mass spectrum before calculating the similarity.
A data processing apparatus for chromatographic mass spectrometry according to any one of claims 1 to 4,
The data processing apparatus for chromatographic mass analysis, wherein the second similarity calculation unit executes a process of removing a peak below a predetermined threshold value for each mass spectrum before calculating the similarity.

Images