JP2005091068A - Method for analyzing colored specimen and analyzer therefor - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for more precisely analyzing a coloring component in a colored specimen causing color development, and to provide an analyzer therefor. <P>SOLUTION: This method for analyzing a colored specimen is characterized by including: (a) a step for preparing two-dimensional chromatograms of time and absorption intensity, respectively, of a colored specimen and a non-colored specimen for each wavelength from three-dimensional chromatogram data on time, wavelengths, and absorption intensity, respectively, of the colored specimen and non-colored specimen acquired by liquid-chromatographically analyzing or gel-permeation chromatographically analyzing the colored specimen and non-colored specimen by using a photodiode array detector; (b) a step for comparing the two-dimensional chromatogram of the colored specimen with that of the non-colored specimen to select a component peak existing only in the two-dimensional chromatogram of the colored specimen; (c) a step for preparing an absorption spectrum of wavelengths and absorption intensity from the three-dimensional chromatogram with respect to the selected component peak; and (d) a step for calculating a color specifying value on the selected component from the absorption spectrum. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

  The present invention relates to a color sample analysis method and an analysis apparatus.

  In the manufacturing process of pharmaceuticals, agricultural chemicals, chemical products, and the like, intermediates and products exhibiting a hue different from the original hue may be obtained due to various causes such as mixing of other components and thermal deterioration. If an intermediate or product that shows a hue different from the original hue is obtained, investigate the component that caused the hue change, and investigate the cause of the component. The result is reflected in the manufacturing process so that an intermediate or product having a hue different from the original hue cannot be obtained again.

  In order to investigate the component that caused the hue change, a sample with a normal hue change (hereinafter referred to as a colored sample) is collected, and the colored sample is subjected to absorbance analysis to obtain an ultraviolet / visible light absorbance spectrum. Compare the UV / visible light absorption spectrum of the sample with the UV / visible light absorption spectrum of the sample showing the original hue (hereinafter referred to as the uncolored sample), and select a wavelength region with a difference in absorption intensity. Calculate the color values of the colored sample and the uncolored sample and use them as basic data for coloring. On the other hand, the colored sample is chromatographed using a photodiode array detector and is a component having absorption in the selected wavelength region. And confirming that the component present only in the colored sample has absorption in the selected wavelength region in the absorption spectrum of wavelength and absorption intensity, Min analyzed to be colored due ingredient, was determined.

  However, this analysis method is based on the premise that the component having absorption in the wavelength region is a color-causing component, and even if the component has absorption in the wavelength region, the component is not colored. If there is no influence or a small component, the cause of coloring will not be investigated, and if there are a plurality of components having absorption in the wavelength range, which component is the cause of coloring. Information on which of these components has the greatest effect on coloring, etc., and the cause of coloring may not be fully investigated. A way to do it was necessary.

  Under these circumstances, the present inventors have intensively studied to develop a method and an analysis apparatus for analyzing a colored component that causes coloring in a colored sample more precisely, and using a photodiode array detector. From the three-dimensional chromatogram data of the time, wavelength and absorption intensity of each of the colored sample and the uncolored sample obtained by liquid chromatography analysis or gel permeation chromatography analysis of the colored sample and the uncolored sample, for each wavelength, Create a two-dimensional chromatogram of the time and absorption intensity of each of the colored sample and the uncolored sample, compare the two-dimensional chromatogram of the colored sample with the two-dimensional chromatogram of the uncolored sample, and compare the two-dimensional chromatogram of the colored sample. The component peak that exists only in the gram is selected, and the three-dimensional chromatogram is selected for the selected component peak. By creating an absorption spectrum of wavelength and absorption intensity from the togram, and calculating the color value of the selected component from the absorption spectrum, when there are a plurality of coloring components that cause coloring in the colored sample. Even in such a case, since the color value of each coloring component can be calculated, the magnitude of the influence of each coloring component on the coloring can be understood, and it has been found that a more precise analysis is possible, leading to the present invention.

That is, the present invention relates to (a) the time, wavelength, and time of each of a colored sample and an uncolored sample obtained by liquid chromatography analysis or gel permeation chromatography analysis of a colored sample and an uncolored sample using a photodiode array detector. From the three-dimensional chromatogram data of the absorption intensity, creating a two-dimensional chromatogram of the time and absorption intensity of each of the colored sample and the non-colored sample for each wavelength;
(B) comparing the two-dimensional chromatogram of the colored sample with the two-dimensional chromatogram of the uncolored sample, and selecting a component peak that exists only in the two-dimensional chromatogram of the colored sample;
(C) creating an absorption spectrum of wavelength and absorption intensity from the three-dimensional chromatogram for the selected component peak;
(D) a method for analyzing a colored sample characterized by including a step of calculating a colorimetric value of the selected component from the absorption spectrum; and (A) a colored sample and a non-colored sample using a photodiode array detector. From the three-dimensional chromatogram of the time, wavelength, and absorption intensity of each colored sample and uncolored sample obtained by liquid chromatography analysis or gel permeation chromatography analysis of the sample, each colored sample and uncolored sample for each wavelength. Two-dimensional chromatogram creation means for creating a two-dimensional chromatogram of the time and absorption intensity of,
(B) Comparing the two-dimensional chromatogram of the colored sample and the two-dimensional chromatogram of the uncolored sample prepared by the two-dimensional chromatogram creating means, A component selection means to select;
(C) Absorption spectrum creation means for creating an absorption spectrum of wavelength and absorption intensity from a three-dimensional chromatogram for the component peak selected by the component selection means;
(D) Colorimetric value calculation means for calculating the colorimetric value of the component peak selected by the component selection means from the absorption spectrum created by the absorption spectrum creation means. An apparatus for analyzing a colored sample is provided.

  According to the present invention, it is possible to perform a more precise analysis regarding the coloring component in the colored sample.

  In the present invention, an uncolored sample refers to a sample that exhibits the original hue of the sample, and a colored sample refers to a sample that exhibits a hue different from that of the uncolored sample. For example, when a sample that is originally colorless and transparent is colored yellow, the colorless and transparent sample is an uncolored sample, and a yellow colored sample is a colored sample. For example, when a sample that originally shows yellow is colored green, the yellow sample is an uncolored sample, and the green sample is a colored sample.

  The method and apparatus for analyzing a colored sample of the present invention will be described with reference to the drawings. A flow chart of the analysis method of the present invention is shown in FIG. 1, and a diagram functionally showing an example of the configuration of the analysis apparatus of the present invention is shown in FIG.

  First, (a) using a photodiode array detector, the retention time, wavelength, and absorption intensity of each of a colored sample and an uncolored sample obtained by liquid chromatography analysis or gel permeation chromatography analysis of a colored sample and an uncolored sample are analyzed. A step of creating a two-dimensional chromatogram of time and absorption intensity of each of the colored sample and the non-colored sample from the three-dimensional chromatogram will be described (S1 in FIG. 1).

  For the liquid chromatography analysis or gel permeation chromatography analysis performed in step (a), a commercially available liquid chromatography apparatus or gel permeation chromatography apparatus equipped with a photodiode array (PDA) detector 1 is usually used. Whether to use a liquid chromatography device or a gel permeation chromatography device may be selected as appropriate, but it is preferable to use a liquid chromatography device that is considered to have higher resolution. Such analysis is performed under conditions that allow separation of components contained in the colored sample and the uncolored sample, and analysis conditions such as the eluent, column, and temperature to be used may be appropriately selected according to the sample.

  The colored sample and the uncolored sample may be analyzed as they are, or may be analyzed after being diluted with a solvent, for example. When the analysis is performed after dilution with a solvent, the dilution rate of the colored sample and that of the non-colored sample are usually substantially the same.

  The photodiode array detector 1 is a detector in which a large number of small photodiodes are linearly arranged, and can measure the light intensity of a plurality of wavelengths at the same time. Therefore, the photodiode array detector 1 is used for coloring. By performing liquid chromatography analysis or gel permeation chromatography analysis on each of the sample and the uncolored sample, it is possible to obtain a three-dimensional chromatogram of time, wavelength and absorption intensity for each sample. The wavelength region to be analyzed is usually in the range of 200 to 800 nm.

  The obtained three-dimensional chromatogram is normally stored in the storage means 7 such as a memory. An example of the obtained three-dimensional chromatogram is shown in FIG.

  The two-dimensional chromatogram creation means 2 cuts out the time data and the absorption intensity data of the colored sample and the non-colored sample from the obtained three-dimensional chromatogram for each wavelength, and creates a data table of the time and absorption intensity. Create a two-dimensional chromatogram based on the data table. As such a two-dimensional chromatogram, a two-dimensional chromatogram as shown in FIG. 5 is created by viewing the obtained three-dimensional chromatogram from a plane including the time axis and the absorption intensity axis and projecting the chromatogram onto the plane. . The created two-dimensional chromatogram may be stored in the storage means 7 such as a memory, for example. If a two-dimensional chromatogram is created, the process proceeds to the next step (b).

  Step (b) compares the two-dimensional chromatogram of the colored sample prepared in step (a) with the two-dimensional chromatogram of the uncolored sample, and determines the component peak that exists only in the two-dimensional chromatogram of the colored sample. In the selection step (S2 in FIG. 1), the two-dimensional chromatogram of the colored sample and the two-dimensional chromatogram of the uncolored sample prepared in step (a) are compared for each wavelength, and the two-dimensional chromatogram of the colored sample is compared. A component peak that exists only in the gram is selected by the component selection means 3.

  As a criterion for selecting a component peak that exists only in the two-dimensional chromatogram of the colored sample, for example, the slope value of the curve of the two-dimensional chromatogram of the colored sample and the uncolored sample is compared, and the curve of the two-dimensional chromatogram of the colored sample is compared. The difference between the slope value of the two-dimensional chromatogram of the uncolored sample and the slope value of the curve of the uncolored sample is calculated, and the peak where the difference of the slope value is a predetermined value or more is determined as the component peak existing only in the two-dimensional chromatogram of the colored sample The method of doing is mentioned. Further, for example, the two-dimensional chromatograms of the colored sample and the non-colored sample may be juxtaposed or overlapped to select a component peak that exists only in the two-dimensional romantogram of the colored sample.

  In this case, the two-dimensional chromatograms at all measurement wavelengths created in step (a) may be compared, but the wavelength or wavelength region of the two-dimensional chromatogram to be compared is determined in advance. Two-dimensional chromatograms of wavelengths or wavelength regions may be compared. Compare the two-dimensional chromatogram of the colored sample and the non-colored sample in a shorter time, and select the component peak that exists only in the colored sample. It is preferable to compare the two-dimensional chromatograms of the determined wavelength or wavelength region.

  Compare the visible light or ultraviolet / visible light absorbance spectrum of a colored sample with the visible light or ultraviolet / visible light absorbance spectrum of a non-colored sample. Since there is a high possibility that a component having absorption is present only in a colored sample, such a wavelength or wavelength region may be set as a wavelength or wavelength region of a two-dimensional chromatogram to be compared in advance. Further, when there are a plurality of wavelengths or wavelength regions having different absorption intensities, for example, an absorption wavelength region having a hue close to the hue of the colored sample may be selected, or a wavelength region having a large difference in absorption intensity may be selected. .

  Information such as the wavelength or wavelength region of the component peak selected in step (b) and the absorption intensity may be stored in the storage means 7 such as a memory. When selection of the component peak existing only in the two-dimensional chromatogram of the colored sample is completed (S3 in FIG. 1), the process proceeds to the next step (c).

  Step (c) is a step (S4 in FIG. 1) for creating absorption spectrum data of wavelength and absorption intensity from the three-dimensional chromatogram for the component peak selected in step (b). Reads out the three-dimensional chromatogram of the component peak from the three-dimensional chromatogram obtained in the step (a) stored in the storage means 7 based on the information of the component peak selected in the step (b). Then, wavelength data and absorption intensity data are cut out from the read three-dimensional chromatogram, a data table of wavelength and absorption intensity is created, and an absorption spectrum is created based on the data table. The created absorption spectrum is usually stored in the storage means 7 such as a memory. If the creation of the absorption spectrum is completed for all the component peaks selected above (S5 in FIG. 1), the process proceeds to the next step (d).

  Step (d) is a step (S6 in FIG. 1) for calculating the color value of the selected component from the absorption spectrum created in step (c), and the absorption spectrum created in step (c). Then, the color specification value calculating means 5 calculates the color specification value. The calculated color value is stored in the storage means 7 such as a memory.

As the color values, for example, L ^* a ^* b ^* color system lightness L ^* and color coordinates a ^* , b ^* (see JIS Z 8729) representing hue / saturation, L ^* u ^* v ^* color specification Color coordinates u ^* , v ^* (see JIS Z 8729) representing the lightness L ^* and hue / saturation of the system, hue H, lightness V and saturation C (see JIS Z 8721) of the Munsell color system It is calculated from the tristimulus values of the XYZ color system based on the 2 ° visual field (see JIS Z 8701) and the tristimulus values of the X ₁₀ Y ₁₀ Z ₁₀ color system based on the 10 ° visual field (see JIS Z 8722). Various color values can be mentioned, and color values of the L ^* a ^* b ^* color system are preferred. The standard light may be any of standard light A, standard light B, standard light C, and standard light D ₆₅ (see JIS Z 8720).

Formulas for calculating the tristimulus values X, Y and Z of the XYZ color system are shown below.
(In the formula, S (λ) represents a value at the wavelength λ of the spectral distribution of standard light, and x (λ), y (λ), and z (λ) are values of color matching functions in the XYZ color system. And τ (λ) represents the spectral reflectance or spectral transmittance of the standard solution.)

Such tristimulus values are usually calculated by converting the absorbance value of the absorption spectrum created in step (c) into transmittance or reflectance. For example, the conversion of the absorbance value into the transmittance is performed based on, for example, the relationship between the absorbance and the transmittance (the following formula (5)).
(In the formula, A represents absorbance and T represents transmittance.)

  In calculating the tristimulus values, correction such as standard white board correction may be performed as necessary.

For example, the color values of the L ^* a ^* b ^* color system are calculated from the tristimulus values X, Y and Z of the XYZ color system calculated by the above formulas (1) to (4) ( 5) to (7) (provided that X / X _n > 0.008856, Y / Y _n > 0.008856 and Z / Z _n > 0.008856).
(In the formula, X _n , Y _n and Z _n represent the tristimulus values of the completely diffuse reflecting surface.)

  By calculating the color value, the hue of the component itself can be grasped as a numerical value. Therefore, by comparing the color value and the color value of the colored sample, the component affects the hue of the colored sample. Therefore, it is possible to make a more precise analysis.

  When the calculation of the color values is completed for all the component peaks selected above (S7 in FIG. 1), the process proceeds to the next step (e).

  Step (e) is a table showing the color values of the colored sample and the uncolored sample calculated from the visible light or ultraviolet / visible light absorbance spectrum data of the colored sample and the uncolored sample, and the components calculated in the step (d). This is a step of comparing the color values (S8 in FIG. 1), thereby determining not only whether the component is a cause of coloring of the colored sample but also whether the influence on the coloring of the colored sample is large or small. can do.

An explanation will be given taking the L ^* a ^* b ^* color system as an example. First, the colored sample L ^*, a ^* and b ^* and the uncolored sample L ^*, compared with the a ^* and b ^*, together with a number is picked up significantly different color values, the color specification value of the colored sample Is compared with the color value of the uncolored sample to determine which value is greater. For example, when the color value of the colored sample is larger than the color value of the uncolored sample, it is defined as + (plus), and when the color value of the colored sample is smaller than the color value of the uncolored sample, − (minus) ).

Then, paying attention to the picked-up color value of L ^* , a ^* and b ^* of the component, comparing the color value of the component with the color value of the uncolored sample, which value is larger? To decide. Also in this case, in the same manner as described above, for example, when the color value of the component is larger than the color value of the non-colored sample, it is defined as + (plus), and the color value of the component is smaller than the color value of the non-colored sample. Let the case be-(minus). The comparison result of the color value of the colored sample and the non-colored sample is compared with the result of comparison of the color value of the component and the non-colored sample, and if they are the same (+ (plus) or-(minus)) If the component is different, it is determined that the component has little influence on the coloration or is not likely to cause the coloration. Furthermore, when there are a plurality of components that are determined to be highly likely to cause coloring, the colorimetric values of the components are compared, and it can be determined that the larger the component, the greater the influence on coloring. .

  In this case, for example, the color values of the colored sample and the uncolored sample calculated from the visible light or ultraviolet / visible light absorbance spectrum data of the colored sample and the uncolored sample are stored in the storage means 7 such as a memory, The comparison means 6 compares the color value of the component calculated in (d) with the color value of the stored colored sample and uncolored sample, and picks up a component that is likely to cause coloring. Also good.

  EXAMPLES Hereinafter, although an Example demonstrates this invention further in detail, this invention is not limited to these Examples.

Example 1
Since N- (3,5-dichlorophenyl) -1,2-dimethylcyclopropane-1,2-dicarboxylic imide, which was usually obtained as white crystals, was obtained as pale brown crystals, the color components were analyzed. .

  First, white crystals of N- (3,5-dichlorophenyl) -1,2-dimethylcyclopropane-1,2-dicarboxylic imide (hereinafter abbreviated as uncolored sample) and N- (3,5-dichlorophenyl). ) -1,2-dimethylcyclopropane-1,2-dicarboxylic acid imide light brown crystals (hereinafter abbreviated as colored samples) were subjected to ultraviolet and visible light absorbance measurements under the following conditions, respectively. The absorbance spectrum shown in Fig. 2 was obtained. From the obtained absorbance spectrum, it was found that there was an absorption peak peculiar to the colored sample at 300 to 500 nm.

<Measurement conditions>
Apparatus: UV / visible spectrophotometer (manufactured by Shimadzu Corporation, UV-2400PC)
Measurement wavelength: 200 to 800 nm
Diluting solvent: acetonitrile Measurement sample concentration: 10 mg / mL

Furthermore, the color value in the L ^* a ^* b ^* color system was calculated from the absorbance spectrum. Table 1 shows the calculated color values and comparison values (difference between the color values of the colored sample and the non-colored sample). The calculation conditions for the color values were as follows.

<Calculation conditions for color values>
Wavelength: 380 to 780 nm
Standard light: Standard light A
Field of view: 2 °

From this result, the b ^* having the largest difference is picked up among the color values of the colored sample and the uncolored sample, and the b ^* of the colored sample is larger than the b ^{* of the} uncolored sample. did.

  The colored sample and the non-colored sample were analyzed under the following conditions using a gel permeation chromatography apparatus equipped with a photodiode array detector to obtain a three-dimensional chromatogram. The obtained three-dimensional chromatogram is shown in FIG. 3 (uncolored sample) and FIG. 4 (colored sample). Data was cut out for each wavelength from the obtained three-dimensional chromatogram, and two-dimensional chromatograms of a colored sample and an uncolored sample were prepared. Among the created two-dimensional chromatograms, two-dimensional chromatograms at wavelengths of 254 nm, 350 nm, 400 nm, and 450 nm are shown in FIG. 5 (uncolored sample) and FIG. 6 (colored sample).

<Analysis conditions>
Column: TSKgel G2000H _HR + TSKgel G1000H _HR + TSKgel G1000H _HR (each φ7.8 mm × 30 cm, 5 μm)
Thermostatic bath temperature: 40 ° C
Mobile phase: Tetrahydrofuran (1 mL / min.)
Detection wavelength: 200 to 800 nm
Measurement sample concentration: 30 mg / mL

  Subsequently, when the two-dimensional chromatogram of the colored sample and the two-dimensional chromatogram of the non-colored sample were compared, it was retained as a component peak existing only in the two-dimensional chromatogram of the colored sample in the wavelength region of 350 to 400 nm. A component for about 21 minutes (hereinafter, this component is abbreviated as RT21 component), a component for about 22 minutes (hereinafter, this component is abbreviated as RT22 component), and a component (hereinafter, about 24 minutes) , This component is abbreviated as RT24 component).

For these three components, the wavelength and absorption intensity data were cut out from each three-dimensional chromatogram, a wavelength and absorption intensity data table was created, and an absorption spectrum was created based on the data table. From the absorption spectrum, the color values in the L ^* a ^* b ^* color system were calculated (standard light A was used for the 2 ° field of view when calculating the color values). A part of the cut-out wavelength and absorption intensity data table created for the RT22 component is shown in FIG. 7, and the absorption spectrum is shown in FIG. The color values of each component are shown in Table 2.

The b ^{* of} each component was compared with the b ^{* of the} uncolored sample, and the results are shown in Table 3 below.

From the comparison results shown in Table 3 and the comparison results shown in Table 1, among the above three components, the RT24 component has little or no effect on the coloring, and the two components, the RT21 component and the RT22 component, are colored. It was found that the RT22 component having a large b ^* has a large influence among them.

Example 2
Since N, N′-dibenzyl-2,3-diaminobutane-2,3-dicarboxylic acid, which was usually obtained as a pale yellowish white solid, was obtained as a pale yellow solid, the coloring component was analyzed.

  First, a pale yellowish white solid of N, N′-dibenzyl-2,3-diaminobutane-2,3-dicarboxylic acid (hereinafter abbreviated as a non-colored sample) and N, N′-dibenzyl-2,3- The pale yellow solid of diaminobutane-2,3-dicarboxylic acid (hereinafter abbreviated as a colored sample) was subjected to ultraviolet and visible light absorbance measurement under the conditions shown below, and the absorbance spectrum shown in FIG. 9 was obtained. . From the obtained absorbance spectrum, it was found that there was an absorption peak peculiar to the colored sample at 300 to 500 nm.

<Measurement conditions>
Apparatus: UV-visible spectrophotometer (manufactured by Shimadzu Corporation, UV-2400PC)
Measurement wavelength: 200 to 800 nm
Diluting solvent: 5% by weight sodium carbonate aqueous solution / methanol (1/1 volume ratio)
Measurement sample concentration: 10 mg / mL

Furthermore, the color value in the L ^* a ^* b ^* color system was calculated from the absorbance spectrum. Table 4 shows the calculated color values and comparison values (difference between the color values of the colored sample and the non-colored sample). The calculation conditions for the color values were as follows.

<Calculation conditions for color values>
Wavelength: 380 to 780 nm
Standard light: Standard light A
Field of view: 2 °

From this result, the b ^* having the largest difference is picked up among the color values of the colored sample and the uncolored sample, and the b ^* of the colored sample is larger than the b ^{* of the} uncolored sample. did.

  The colored sample and the non-colored sample were analyzed using a liquid chromatography apparatus equipped with a photodiode array detector under the following conditions to obtain a three-dimensional chromatogram. Data was cut out for each wavelength from the obtained three-dimensional chromatogram, and two-dimensional chromatograms of a colored sample and an uncolored sample were prepared. Among the created two-dimensional chromatograms, two-dimensional chromatograms having wavelengths of 254 nm, 400 nm, and 450 nm are shown in FIG. 10 (uncolored sample) and FIG. 11 (colored sample).

<Analysis conditions>
Column: SUMPAX ODS A212 (φ6mm × 150mm, 5μm)
Thermostatic bath temperature: 40 ° C
Mobile phase: Liquid A 1.4% by volume acetic acid aqueous solution Liquid B Methanol
10% B solution / 90% A solution-(45 minutes) → 100% B solution (hold 15 minutes)
Detection wavelength: 200 to 800 nm
Flow rate: 1 mL / min Measurement sample concentration: 10 mg / mL

  Subsequently, when the two-dimensional chromatogram of the prepared colored sample and the two-dimensional chromatogram of the non-colored sample were compared, a retention time of about 37 was obtained as a component peak existing only in the two-dimensional chromatogram of the colored sample at a wavelength of 400 nm. Component (hereinafter, this component is abbreviated as RT37 component), component having a retention time of about 41 minutes (hereinafter, this component is abbreviated as RT41 component), and component having a retention time of about 43 minutes (hereinafter, this component) Is abbreviated as RT43 component).

For these three components, the wavelength and absorption intensity data were cut out from each three-dimensional chromatogram, a wavelength and absorption intensity data table was created, and an absorption spectrum was created based on the data table. From the absorption spectrum, the color values in the L ^* a ^* b ^* color system were calculated (in the calculation of the color values, standard light A was used and a 2 ° field of view was used). A part of the cut-out wavelength and absorption intensity data table created for the RT43 component is shown in FIG. 12, and the absorption spectrum is shown in FIG. The color values of each component are shown in Table 5.

The b ^{* of} each component was compared with the b ^{* of the} uncolored sample, and the results are shown in Table 6 below.

From the comparison results shown in Table 6 and the comparison results shown in Table 4, all of the above three components have an effect on coloring, and among them, the RT43 component having the largest b ^* has the most influence on the coloring. It was found that the RT41 component then affected the coloration.

It is a flowchart of the analysis method of this invention. It is an absorbance spectrum of the uncolored sample and colored sample of Example 1. 3 is a three-dimensional chromatogram of an uncolored sample of Example 1. FIG. 3 is a three-dimensional chromatogram of a colored sample of Example 1. FIG. 2 is a two-dimensional chromatogram of retention time and absorption intensity of an uncolored sample of Example 1. FIG. 2 is a two-dimensional chromatogram of retention time and absorption intensity of the colored sample of Example 1. FIG. It is a part of data table which extracted the data of wavelength and absorption intensity from the three-dimensional chromatogram of RT22 component of Example 1. 4 is an absorption spectrum of RT22 component of Example 1. It is an absorbance spectrum of an uncolored sample and a colored sample of Example 2. It is a two-dimensional chromatogram of the retention time and absorption intensity of the uncolored sample of Example 2. It is a two-dimensional chromatogram of the retention time and absorption intensity of the colored sample of Example 2. It is a part of data table which extracted the data of wavelength and absorption intensity from the three-dimensional chromatogram of RT43 component of Example 2. It is an absorption spectrum of RT43 component of Example 2. It is the figure which showed the structure of an example of the analyzer of this invention functionally.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Photodiode array detector 2 Two-dimensional chromatogram preparation means 3 Component selection means 4 Absorption spectrum preparation means 5 Colorimetric value calculation means 6 Comparison means 7 Storage means

Claims (5)

(A) Three-dimensional time, wavelength, and absorption intensity of each of a colored sample and an uncolored sample obtained by liquid chromatography analysis or gel permeation chromatography analysis of a colored sample and an uncolored sample using a photodiode array detector Creating a two-dimensional chromatogram of the time and absorption intensity of each of the colored and uncolored samples from the chromatogram data for each wavelength;
(B) comparing the two-dimensional chromatogram of the colored sample with the two-dimensional chromatogram of the uncolored sample, and selecting a component peak that exists only in the two-dimensional chromatogram of the colored sample;
(C) creating an absorption spectrum of wavelength and absorption intensity from the three-dimensional chromatogram for the selected component peak;
(D) A method for analyzing a colored sample, comprising: calculating a colorimetric value of the selected component from the absorption spectrum. Step (b)
(B ′) Obtaining visible light or ultraviolet / visible light absorbance spectra of a colored sample and an uncolored sample, comparing the spectra, selecting a wavelength region having a difference in absorption intensity, and selecting the wavelength region in the selected wavelength region The step of comparing the two-dimensional chromatograms of the colored sample and the non-colored sample obtained in (a) and selecting the component peak existing only in the two-dimensional chromatogram of the colored sample. Analysis method. (E) Comparison of the color values of the colored sample and the non-colored sample calculated from the visible light or ultraviolet / visible light absorption spectra of the colored sample and the non-colored sample with the color values of the components calculated in (d) above. The method for analyzing a colored sample according to claim 1, further comprising the step of: (A) Three-dimensional time, wavelength, and absorption intensity of each of a colored sample and an uncolored sample obtained by liquid chromatography analysis or gel permeation chromatography analysis of a colored sample and an uncolored sample using a photodiode array detector A two-dimensional chromatogram creation means for creating a two-dimensional chromatogram of each time and absorption intensity of a colored sample and an uncolored sample for each wavelength from the chromatogram,
(B) Comparing the two-dimensional chromatogram of the colored sample and the two-dimensional chromatogram of the uncolored sample prepared by the two-dimensional chromatogram creating means, A component selection means to select;
(C) Absorption spectrum creation means for creating an absorption spectrum of wavelength and absorption intensity from a three-dimensional chromatogram for the component peak selected by the component selection means;
(D) Colorimetric value calculation means for calculating the colorimetric value of the component peak selected by the component selection means from the absorption spectrum created by the absorption spectrum creation means. Colored sample analyzer. (E) Comparing means for comparing the color value of the colored sample and the non-colored sample calculated from the visible light or ultraviolet / visible light absorption spectrum of the colored sample and the non-colored sample with the color value of the component peak is further provided. The colored sample analyzer according to claim 4.