JP3524968B2 - Determination of calcium oxalate monohydrate and calcium oxalate dihydrate in living stones by infrared spectroscopy - Google Patents

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a quantitative analysis of calcium oxalate monohydrate and calcium oxalate dihydrate in a urinary calculus by infrared spectroscopy. In particular, substantially 78 by infrared spectroscopy
This gives a quantitative method focusing on the absorption band at 0 cm ^-1 .

[0002]

2. Description of the Related Art Conventionally, X-ray scattering method and thermal analysis method have been used as a method for quantitatively analyzing urinary calculi. However, these methods are not always convenient because they require special equipment. Also, X
The method using lines has a problem in rapid measurement because it requires more skill in preparing a measurement sample and it takes time to analyze the obtained data. On the other hand, the analysis of urinary tract stones using infrared spectroscopy has been widely used for its qualitative analysis. Several attempts to quantitatively analyze calcium monohydrate (COM) and calcium oxalate dihydrate (COD) have been reported so far (Takasaki, E.). , Url. Int. (Urol. Int.), 30; 228 (1975); Martin, X., etc., Kidney int., 25; 948 (1984); Oka, tea.
(Oka, T.), J. Url Oel. (J. Urol.),
134; 813 (1985); Moriguchi, H. (Moriguchi,
H.), JPN. J. Url Oel. (J
pn. J. Urol.), 77; 1485 (1986)), but the fact is that a satisfactory method has not been developed. In such a conventional quantitative method, 600 to 610 cm ^-1 , 780 cm ^-1 ,
An attempt was made to measure calcium oxalate monohydrate and calcium oxalate dihydrate in urinary calculi using infrared absorption at 920 cm ⁻¹ or 1320 cm ⁻¹ .

It has been reported that calcium oxalate (CaOx) and calcium phosphate (CaP) are major constituents (60 to 80%) of urinary calculi. Magnesium / ammonium phosphate stones or uric acid stones are not so often observed. Such uric acid stones contain, in addition to the above components, calcium urate, calcium carbonate, sodium acid urate, ammonium acid urate, cystine, xanthine, protein, magnesium monohydrogen phosphate, calcium monohydrogen phosphate, etc. at various ratios. It may have a complicated composition.

If the urinary calculus is composed of a single component, it is easy to analyze the urinary calculus by infrared absorption, but the urinary calculus is usually composed of two or more components. It is quite difficult to analyze urinary calculi by infrared absorption because they are mixed in the urine. Almost all calcium oxalate uroliths are composed of at least three or more of calcium oxalate monohydrate, calcium oxalate dihydrate, calcium phosphate (CaP) and other ingredients. Made of By the way, these calcium oxalate urinary calculi are calcium oxalate monohydrate (COM) urinary calculi depending on the amount of calcium oxalate monohydrate and calcium oxalate dihydrate contained therein. Calcium oxalate / dihydrate (COD) urinary stones are classified into two categories, but calcium oxalate / monohydrate is clinically prone to the formation of calcium oxalate / dihydrate urinary stones. It has been pointed out that urinary calculi formation is more likely to recur than in patients with urolithiasis.

Therefore, since a more strict follow-up is required for a patient with calcium oxalate dihydrate urinary calculi, the calcium oxalate monohydrate contained in the calcium oxalate urinary calculus is accurately determined. It is required to quantify the amount of calcium oxalate dihydrate.
Furthermore, in clinical practice, it is required to determine the amount of calcium oxalate / monohydrate and calcium oxalate / dihydrate contained in the stones quickly and accurately, especially in stones obtained from urine. Has been. This is because calculi are crushed by shock waves and treated, but since dihydrate calculi are larger than monohydrate calculi and their hardness is different, shock waves during treatment etc. Quantitative analysis is of great significance in the treatment of lithotripsy, such as the control of, and rapid and accurate quantification is required.

[0006]

According to infrared spectroscopic analysis,
Attempts to quantify the amounts of calcium oxalate / monohydrate and calcium oxalate / dihydrate contained in urinary calculi were carried out by using, for example, Oka and the like from the absorbance ratio of the absorption bands at 660 cm ⁻¹ and 610 cm ⁻¹ to determine oxalic acid. A method for obtaining the content rate of calcium dihydrate has been published (Oka, Tea.
T.), J. URL. (J. Urol.), 134;
813 (1985)). However, with this method, it is necessary to solve the complex quadratic function relationship, and the accuracy is not very good, or the absorption band used overlaps the absorption spectrum of calcium phosphate, and the measurement of stones containing calcium phosphate that is very general There are problems such as becoming difficult. Such problems are also pointed out by Moriguchi et al. In the conventional quantification method by infrared spectroscopic analysis, there is a problem that a complicated operation of weighing 1 mg of a calculus sample is required to calculate the content.

Moriguchi et al. Investigated the infrared spectrum of a calculus sample in an attempt to solve these problems and found that it was 920 cm.
The quadratic equation from the absorption band of ^-1 to seek a relation between the content of the depth and oxalate calcium dihydrate of the absorption band, further calcium oxalate content in stones, 1,320Cm ^{- 1} , 1,100 to 1,000 cm ^-1
Calculated separately from the ratio of the absorption band depths. However, with this method, the content of calcium oxalate / dihydrate is 35%.
It is pointed out that the absorption band at 920 cm ^-1 becomes unclear below. In other words, it is pointed out that there is a problem that the ratio of calcium oxalate / monohydrate to calcium oxalate / dihydrate cannot be determined in a stone sample having a low calcium oxalate / dihydrate content.

Moriguchi et al., 660 cm ^-1 and 610 cm
Considering to determine the content of calcium oxalate / dihydrate from the absorbance ratio in the absorption band of ^-1 shows that the content of calcium oxalate / dihydrate is significantly higher than the actual content. It points out the problem of going out. Moriguchi,
Absorption band depth ratio of 1,320 cm ^-1 , 1,100 ~
The ratio of the absorption band of the depth of 1,000 cm ^-1 and 780 cm ^-1
Consider the depth ratio of the absorption band of the correlation between the calcium phosphate content / calcium oxalate content, obtains the correlation between the content of calcium oxalate monohydrate, respectively the absorption band of 780 cm ^-1 Depth ratio and 1,320c
It has been reported that there is a quadratic relationship with the ratio of the absorption band depth of m ^-1 (Hideo Moriguchi et al., "Urine Bulletin", 37: 1-5, 1991).

As described above, in order to quantify calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi by infrared spectroscopic analysis, it is necessary to solve a complicated quadratic equation. , Calcium oxalate monohydrate and calcium oxalate dihydrate in the stone sample with low calcium oxalate dihydrate content, which is affected by the calcium phosphate content in urinary calculi. There were problems such as not being able to decide.

[0010]

The present inventors have been able to quantitatively analyze calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi by infrared spectroscopic analysis, which has clinical significance. If we proceed with research on a method that can obtain certain data easily and accurately, and pay attention to a specific absorption band of the spectrum obtained by infrared spectroscopic analysis, calcium oxalate monohydrate (COM) and calcium oxalate can be found.・
The present invention has been completed by finding that the proportion of dihydrate (COD) can be accurately determined by simply relating it to its infrared absorption spectrum.

That is, according to the present invention, in the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopic analysis, substantially 8
Using the absorption band at 00 cm ^-1, the content ratio of calcium oxalate monohydrate and calcium oxalate dihydrate was related to the measured infrared absorption spectrum, and based on this relationship, oxalic acid in urinary calculi This is a method characterized by quantifying calcium / monohydrate and calcium oxalate / dihydrate. So far, substantially 800 cm ^-1
There is no report that quantified calcium oxalate monohydrate and calcium oxalate dihydrate using infrared absorption spectrum in.

From the urinary tract stone sample, a substantially pure calcium oxalate monohydrate calculus sample and a substantially pure calcium oxalate dihydrate calculus sample were prepared, respectively. The infrared absorption spectrum of the standard calculus sample was measured by an infrared spectrometer, preferably a Fourier transform infrared spectrometer (FT-IR), and calcium oxalate / monohydrate was obtained from the obtained infrared absorption spectrum. An absorption band based on a halide and calcium oxalate dihydrate, which is effective for quantitative analysis is selected. As a result of the measurement, 1620 cm ^-1 and 1320
By using the absorption band at cm ⁻¹ , there is little difference in the shape between the absorption spectrum of calcium oxalate / monohydrate and the absorption spectrum of calcium oxalate / dihydrate, and highly accurate measurement is possible. Determined to be difficult.
Also, at 950 cm ^-1 , 920 cm ^-1 and 890 cm ^-1 , the respective absorption band peaks are low, and it is difficult to measure trace amounts with high accuracy, and it is also difficult to measure in the presence of other calcium phosphates. Is determined.

It has been pointed out that there is a considerable difference in the infrared absorption spectrum between the calculus sample and the chemical reagent even if the same calcium oxalate / hydrate is used. It is considered that this is due to the influence of organic substances such as proteins contained in the calculus sample. Therefore, when analyzing the band effective for quantification by infrared spectroscopy, calcium oxalate / hydrate from the calculus sample should be analyzed. A sample must be used. As a result of the analysis, it was found that the use of the absorption band at 780 cm ⁻¹ shows that the absorption peaks of calcium oxalate / monohydrate and calcium oxalate / dihydrate are significantly different and It is judged to be effective because it is sharp. This substantially 7
In order to utilize the infrared absorption spectrum appearing at 80 cm ⁻¹ , the infrared absorption spectrum appearing in the range of 700 to 900 cm ⁻¹ including the absorption band was analyzed.
By using the value of the infrared absorption spectrum in substantially wave number of 800 cm ^-1 in the vicinity of the absorption peak of the 0 cm ^-1, and the content of calcium oxalate monohydrate and calcium oxalate-dihydrate in stone A simple correlation can be derived between.

According to the present invention, this correlation is a linear equation (linear equation), which is substantially the same as a calculus sample composed of substantially pure calcium oxalate monohydrate prepared from a urinary calculus sample. Pure calcium oxalate
A calibration curve based on each infrared absorption spectrum was prepared using a stone sample made of dihydrate and a standard sample obtained by mixing them at an arbitrary ratio, and calcium oxalate
A correlation according to a linear equation can be obtained between the content ratio between the monohydrate and calcium oxalate / dihydrate and the value of the infrared absorption spectrum. According to this relational expression, it is possible to easily quantify calcium oxalate / monohydrate and calcium oxalate / dihydrate by simply using the value of the infrared absorption spectrum. In the present invention, in particular, the ratio of the concentration of calcium oxalate / monohydrate (COM) / calcium oxalate / dihydrate (COD) ([COM] / [COD])
Between the ratio of infrared absorption in the specific absorption band and a ratio according to a linear equation, or calcium oxalate dihydrate (COD) / calcium oxalate monohydrate Compound (COM) concentration ratio ([COD] /
[COM]] and the ratio of infrared absorption in the specific absorption band, a relationship according to a linear equation (linear equation) is obtained.

According to one embodiment of the present invention, a baseline connecting the bottoms of the valleys of the spectrum on both sides of the peak of the infrared absorption spectrum substantially appearing at 780 cm ^-1 is set, 7 based on the baseline
By using infrared absorption spectrum in substantially wave number of 800 cm ^-1 in the vicinity of the peak of the peak and the absorption of the absorption of 80 cm ^-1, the calcium oxalate monohydrate / oxalate calcium dihydrate Concentration ratio and infrared absorption ratio of calcium oxalate / monohydrate / calcium oxalate / dihydrate based on the baseline, or of calcium oxalate / dihydrate / calcium oxalate / monohydrate Between the concentration ratio and the infrared absorption ratio of calcium oxalate / dihydrate / calcium oxalate / monohydrate with respect to the baseline, there is a relationship according to a linear equation (linear equation). Based on this relationship, quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi by infrared spectroscopy can be performed. The infrared absorption peaks at 780 cm ⁻¹ and 800 cm ⁻¹ are substantially the 780 cm ⁻¹ and 800 c ⁻¹ , respectively.
If it can be regarded as an infrared absorption peak of m ^-1 , it can be regarded as an infrared absorption peak intended for use in the present invention. Similarly, it is understood that the same applies to the valleys of the spectra on both sides of the peak, and the wave number near the absorption peak is not limited as long as the method of the present invention can be substantially applied.

The concept of the invention is illustrated by illustrating one specific embodiment according to the invention. Prepare calcium oxalate monohydrate standard sample and calcium oxalate dihydrate standard sample from at least 200 urinary calculi.
These standard samples are each almost 100% pure even when subjected to X-ray scattering analysis. The COM standard sample and the COD standard sample are stored at 4 ° C or lower until use, and the standard sample is stored as calcium oxalate / monohydrate /
Weight ratio of calcium oxalate / dihydrate (COM / CO
D weight ratio) is 0 to 3.906, and further, the weight ratio of calcium oxalate / dihydrate / calcium oxalate / monohydrate (COD / COM weight ratio) is 0 to 1.050. Mix just prior to infrared spectroscopy. 2 mg or 3 mg of each powder sample was mixed with potassium bromide powder or potassium chloride powder, and tablets (pellets, pel)
Let). The measuring method can be appropriately selected from among the methods suitable for infrared spectroscopic analysis.

An infrared spectrometer can be used to obtain the infrared absorption spectra of these tablet samples. Examples of the infrared spectrometer include a dispersive infrared spectrometer and a Fourier transform infrared spectrometer. A Fourier transform infrared spectrometer can be preferably used. Fourier transform infrared spectrometers include, for example, Bio-Rad, Bruker, Perkin-E.
Lmer, Shimadzu, JEOL, JASCO, Fuji Electric, Hitachi, etc. are sold. For example, 4000-
A Fourier transform infrared spectrometer with a measurable range of 400 cm ^-1 can be used. 1A shows a pure calcium oxalate monohydrate standard sample, and FIG. 1B shows a pure calcium oxalate dihydrate standard sample. C of FIG. 1 shows an infrared transmission spectrum of calcium phosphate.

According to the respective measurements, the infrared absorption spectra of calcium oxalate / monohydrate, calcium oxalate / dihydrate, calcium phosphate (CaP), etc. have infrared absorptions at their respective wave numbers. To be observed. C
In OM 670cm ^-1, 890cm ^-1, and 950cm
^-1 bands were observed, while COD was 610c
Bands at m ⁻¹ and 920 cm ⁻¹ are observed. 78
0cm each band of ^-1 and 1320 cm ^-1 is COM and CO
Observed in both D. 560 for CaP
cm ^-1, 600cm ^-1, and each band of 1020 cm ^-1 is observed. Of the urinary tract stones, the combination of calcium oxalate (CaOx) and calcium phosphate (CaP) is most often observed. It can be said that the infrared absorption spectrum of CaP has a great influence on the infrared absorption spectrum of urinary calculus CaUx, and from the observation that it has a large influence particularly in the range of 600 to 610 cm ^−1. be able to. Therefore, 600-6
If a band in the range of 10 cm ^-1 is used, COM and C
It will be appreciated that it is difficult to perform a quantitative analysis of COM and COD by analyzing common uroliths that normally contain CaP in addition to OD.

890 cm ^-1 , 920 cm ^-1 , and 950
infrared absorption in cm ^-1 is observed that the absorption than the infrared absorption at 780cm ^-1 is weak. So C
It can be seen that it is better to pay attention to the infrared absorption at 780 cm ^{-1, which} is not affected by aP. Therefore, we examined the infrared absorption peak and the absorption valley in this region. FIG. 2 shows a comparison of absorption curves of calcium oxalate monohydrate and calcium oxalate dihydrate in the range of 700 to 900 cm ⁻¹ . The change in infrared absorption is CO
D is small and COM is large. 780-900 cm
^In the range of ^-1 , there is a difference in the spectrum between calcium oxalate monohydrate and calcium oxalate dihydrate. COM shows a sharp gradient in the range of 780 to 820 cm ^-1 , while COD is 780 to 90.
It can be confirmed that the gradient is gentle in the range of 0 cm ^-1 . The difference in absorption between the pure COM and pure COD samples varied from 780 cm ⁻¹ with increasing wavenumber,
The difference in absorption between the COM and COD in 80 cm ^-1 vicinity of 800 cm ^-1 is observed that maximized. 2C
The result of plotting the absorbance difference | ab | between OD and COM in steps of 5 cm ⁻¹ is shown in FIG. 3.

First, in FIG. 6, a line connecting valleys O and Q on both sides of the peak at 780 cm ^-1 is defined as a base line c, a point on the base line c is represented by C, and a peak apex is represented by P. The point at 800 cm ⁻¹ on the baseline c is designated as C ′. A line CP is drawn between the C at 780 cm ⁻¹ and the absorption peak P, and further the line C−
Draw a line C'-P 'parallel to P and through the C'at 800 cm ^-1 . The distance from C to P is | C-
Indicated by P |, | C-P | is equal to | C'-P '|. Next, the point where the line C′-P ′ at 800 cm ⁻¹ intersects with the observed absorption spectrum is defined as R ′. Then, a line R'-R parallel to the line c (C'-C) is drawn. R
Is the point on line CP at 780 cm ^-1 .
The line P'-P is also parallel to the line c (C'-C).

Similarly, as shown in FIG. 4, R ″ corresponding to R ′ is obtained for a pure calcium oxalate monohydrate (COM) standard sample, and as shown in FIG. 5, pure calcium oxalate · R'for dihydrate (COD) standard sample
R ″ ′ corresponding to 800 as shown in FIG.
The difference in absorption between the COM standard calculus sample and COD standard calculus sample in cm ^-1 is taken into account that it is observed that maximized, Looking at the absorption band of 800 cm ^-1, R ''
Is located near C'and only a short distance from C ', while R'"is also located near P'and a short distance from P '.

Thus, the position of R'at 800 cm ^-1 can be considered to be influenced by the ratio of COM / COD concentrations. If the COM / COD concentration ratio increases, the position of R'will be located closer to R "or C '. On the other hand, when the COM / COD concentration ratio decreases, the position of R ′ comes closer to R ′ ″ or P ′. On the other hand, the infrared absorption observed for the urinary stone sample at 780 cm ^-1 is
It is considered to include absorption by M and absorption by COD. Then, since the lines (C'-C) and (P'-P) are parallel, | P-C | is equal to | P'-C '|. Furthermore, | C'-R '| is the absorption of calcium oxalate dihydrate (COD), and | P'-R' | is the absorption of calcium oxalate monohydrate (COM). I suppose. Then, similarly, the lines (C'-C), (R '
-R) and (P'-P) are parallel, | C'-R '
| Is equal to | C-R | and | P'-R '| is | P-
It is equal to R |.

Thus, | CR | can be the absorption of COD, and | PR | can be the absorption of COM. | C'-P '| at 800 cm ^-1 is 78
Equal to | CP | at 0 cm ⁻¹ , COM and CO
It is assumed that this corresponds to the sum of absorptions of D ([COM + COD]). Then, the COD absorption ([COD]) is calculated as | C'-
When R ′ |, that is, | C−R |, the absorption becomes [COM + COD] = [COM] + [COD], and [COM] = [COM + COD] − [COD], so [COM] = | P ′ −R ′ | = | P−R |

If the ratio of absorption of COM / COD is proportional to the ratio of concentration of COM / COD as predicted by Lambert-Beer's law, the infrared absorption spectrum at 800 cm ^-1 ｜
C'-R '| (= [COD]), that is, | C-R |
Infrared absorption spectrum at 80 cm ^-1 | P-C |
It is predicted that the ratio of COM / COD concentrations can be obtained using ([COM + COD]). There 80
The concentration was compared with the measured value using the absorption band at 0 cm ⁻¹ . The relationship was examined by actually measuring the standard stone sample prepared with the above-mentioned mixing ratio. 800
The absorption band at cm ⁻¹ is easy to read. 800
Although it is possible to perform quantitative measurement by infrared spectroscopy using bands other than cm ^-1 , it is substantially 800 cm
^It is preferred to use an absorption band of ^-1 . In actual use, | C′−R ′ | and | P′−R ′ | may be replaced with | C−R | or | P−R |. | CR | can be the absorption of COD, and | P
-R | may be the absorption of COM.

Absorption ratio [COM] / [COD] (Y)
Is Y = | P'-R '| / | C'-R' | (= | P-R | /
It can be seen that it can be represented by | C'-R '|). Concentration ratio COM / C
As a result of the measurement, the following cubic equation is established between the ratio of the infrared absorption of COM / COD and the ratio of the concentration of COM / COD when X> 0, where OD is X (Fig. 7). The ratio of COM and COD concentrations can be expressed as a percentage. ^{Y = 0.163X 3 -1.484X 2 + 4.778X} +
0.145

However, when 0 <X <1, COM / C
It is recognized that the following linear relationship holds between the infrared absorption ratio of OD and the COM / COD concentration ratio (FIG. 8). Y = 3.376X + 0.350 Similarly, the infrared absorption ratio Y'of COD / COM and CO
It is recognized that the following linear relationship holds with the D / COM concentration ratio X ′ (FIG. 9). Y '= 0.143 X' + 0.129 This relationship holds when 0 <X '<1. This linear relationship is the ratio of infrared absorption of COD / COM to COD / COM.
COM / COD as observed between the concentration ratio of
It is also observed between the infrared absorption ratio and the COM / COD concentration ratio.

In the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi by infrared spectroscopy, the absorption band at 800 cm ^-1 was examined. Is the infrared absorption of COD, and the absorption band at substantially 780 cm ⁻¹ is the infrared absorption of the sum of COM and COD, and (1) the concentration ratio COM / COD is X, and the absorption ratio [COM] / [CO
D] is Y and 0 <X <1, the following linear relational expression: Y = aX + b a = 2.720 to 3.689 b = 0 to 1.33
0

(2) Infrared absorption ratio [COD] / [CO
M] is Y ′, the COD / COM concentration ratio is X ′, and 0 <X ′ <1, the following linear relational expression: Y ′ = a′X ′ + b ′ a ′ = 0.130 ~ 0.170 b '= 0.080
It is confirmed that ~ 0.152 can be used to quantitatively analyze calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary tract stones. It is obvious that the calculus is not particularly limited as long as it occurs in the living body, and it is particularly used for measuring urinary tract stones. Living stones include urinary tract stones such as kidney stones, bladder stones, ureteral stones, and urethral stones,
Examples include gallstones, blood stones, and pancreatic stones.

Therefore, in the present invention, in the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopy, an infrared absorption spectrum substantially appearing at 780 cm ^-1 is obtained. 900 cm ⁻¹ on the high wavenumber side near the absorption peak and the absorption peak.
To obtain the relationship between the amounts of calcium oxalate / monohydrate and calcium oxalate / dihydrate and the infrared absorption spectrum according to a linear equation (linear equation). The present invention provides a method characterized by quantifying calcium oxalate / monohydrate and calcium oxalate / dihydrate in a living stone based on this relationship.

The present invention also provides a substantially quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopy for a total of 780c.
The infrared absorption spectrum peak appearing at m ^-1 is sandwiched, and a baseline is set that connects substantially the bottoms of the valleys of the spectrum on both sides of the peak, and the baseline, the absorption peak, and the vicinity of the absorption peak. 900c on the high wavenumber side of
Using the band of infrared absorption spectrum up to m ^−1, the ratio of the concentration of calcium oxalate / monohydrate to the concentration of calcium oxalate / dihydrate and the calcium oxalate / monohydrate based on the baseline The relationship between the calcium oxide and the ratio of infrared absorption of calcium oxalate / dihydrate according to the linear equation was obtained, and based on this relationship, calcium oxalate / monohydrate and oxalate in living stones were obtained. It is a method characterized by quantifying calcium acid dihydrate. In this method, in particular, the wave number near the absorption peak is substantially 800 cm ⁻¹ .

In summary, the present invention is substantially equivalent to 8 in the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopy.
Using the absorption band at 00 cm ^-1, the content ratio of calcium oxalate / monohydrate and calcium oxalate / dihydrate was related to the measured infrared absorption, and based on this relationship, calcium oxalate in urinary calculi・ This method is characterized by quantifying monohydrate and calcium oxalate / dihydrate.

In a more specific constitution, the present invention provides the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopy.
The infrared absorption spectrum peak appearing substantially at 780 cm ⁻¹ and the infrared absorption spectrum appearing substantially at 800 cm ⁻¹ are used, and the 780 cm ⁻¹ based on the baseline of the peak of the infrared absorption spectrum appearing at 780 cm ^−1. Assuming a parallel homology between ^-1 and 800 cm ^-1 , 800c
The infrared absorption spectrum appearing at m ^-1 is assumed to be the amount of infrared absorption of calcium oxalate dihydrate, and
^-1 shows the infrared absorption spectrum of calcium oxalate
As the sum of infrared absorptions of monohydrate and calcium oxalate / dihydrate, the content ratio of calcium oxalate / monohydrate and calcium oxalate / dihydrate in the sample and the 780c
The relationship between the measured infrared absorption spectrum appearing at m ⁻¹ and the measured infrared absorption spectrum is obtained according to a linear equation, and based on this relationship, calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones It is a method characterized by quantifying a compound.

In a typical embodiment, the present invention provides a quantitative analysis of calcium oxalate monohydrate and calcium oxalate dihydrate in urinary stones by infrared spectroscopy, which is substantially 800 cm ^-1. Absorption band at COD
And the absorption band at substantially 780 cm ⁻¹ is the infrared absorption of the sum of COM and COD, and (1) the concentration ratio COM / COD is X, and the absorption ratio CO
When M / COD is Y and 0 <X <1, the following linear relational expression: Y = aX + b a = 2.720 to 3.689 b = 0 to 1.33
0 (2) The infrared absorption ratio of COD / COM is Y ', and CO
When the ratio of the D / COM concentrations is X ′ and 0 <X ′ <1, the following linear relational expression: Y ′ = a′X ′ + b ′ a ′ = 0.130 to 0.170 b ′ = 0.080
The present invention relates to a method for quantitatively analyzing calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary tract stones by utilizing ~ 0.152. In this method, it is particularly preferable that substantially a = 3.376, b = 0.350, a ′.
A method is provided in which = 0.143 and b '= 0.129.

Various modifications are possible as long as the basic concept of the present invention is utilized, and all of these modifications are also included in the present invention. Further, the present invention provides a substantially quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi by infrared spectroscopic analysis.
Utilizing the absorption band of the 0 cm ^-1 and 800 cm ^-1 method of performing determination of calcium oxalate monohydrate and calcium oxalate-dihydrate are given. As described above, the present invention, in the quantitative analysis by infrared spectroscopic analysis of calcium oxalate monohydrate and calcium oxalate dihydrate in urinary calculi, substantially calcium oxalate monohydrate A linear relational expression was found between the ratio of the concentration of calcium oxalate / dihydrate and the bands of the two absorption spectra, and based on the linear relational expression, the relationship between calcium oxalate / monohydrate in urinary calculi and A method for quantitative analysis of calcium oxalate dihydrate is provided.

The stone samples to be measured can be substantially in all ranges with respect to the ratio of calcium oxalate monohydrate to calcium oxalate dihydrate. The calculus sample is not particularly limited as long as it is a calcium oxalate calculus, but as is apparent from the above, a sample containing calcium phosphate can also be measured. The calculus sample can usually be made into a powder, mixed with potassium bromide, etc., and made into a tablet for convenience of measurement, and then subjected to the measurement, but there is no problem in the purpose because of the method commonly used in the field. All methods are available. The results can also be obtained at the same time as the measurement by incorporating a calculation formula or a program related thereto into a computer or the like of the infrared spectrometer.

[0036]

EXAMPLES The present invention will now be described in more detail with reference to examples, but the present invention is not limited to these examples. Example (1) Preparation of calcium oxalate / monohydrate standard sample and calcium oxalate / dihydrate standard sample from urinary calculi From at least 200 urinary calculi, calcium oxalate / monohydrate standard sample and oxalic acid A calcium dihydrate standard sample was prepared. Calcium oxalate monohydrate standard calculus samples are Weddell scores
A stone sample with 0 ite score) was collected and prepared. The calcium oxalate dihydrate standard stone sample was prepared by collecting a stone sample having a Wederite score of 8 points. When these standard samples were subjected to X-ray scattering analysis, it was confirmed that each of them had a purity of almost 100%. The COM standard sample and the COD standard sample were stored at 4 ° C. or lower until use. Use this standard sample as calcium oxalate monohydrate / calcium oxalate
The weight ratio of the dihydrate (COM / COD weight ratio) is 0 to
In the range of 3.906, the weight ratio of calcium oxalate / dihydrate / calcium oxalate / monohydrate (COD
/ COM weight ratio), the mixture was mixed in the range of 0 to 1.050 immediately before the infrared spectroscopic analysis. About 2 mg or about 3 mg of each powder sample is about 200 mg of potassium bromide powder (KBr for spectroscopic analysis, manufactured by Merck).
It was mixed with and molded into tablets (pellets). In preparing the powder, the powder was pulverized so as not to cause metamorphosis and to be uniform.

(2) Analysis of infrared absorption spectrum Fourier transform infrared spectrometer (FT-IR) having a measurable range of 4000 to 400 cm ^-1 (model VALOR-I)
II, manufactured by JASCO Corporation) was used to analyze these tablet samples. The measurement conditions are 16 times of integration and gain is A
uto, decomposition 4 cm ^-1 , apotization CS,
The diameter of the apatcher was 8 mm. The obtained results are shown in FIG. 1A shows a pure calcium oxalate monohydrate standard sample, and FIG. 1B shows a pure calcium oxalate dihydrate standard sample. C of FIG. 1 shows an infrared transmission spectrum of calcium phosphate. Next, the peak of infrared absorption and the valley of absorption were examined. According to each measurement, COM, COD, calcium phosphate (Ca
In the infrared absorption spectra of P) and the like, infrared absorptions at characteristic wave numbers were observed. 670 cm at COM
^-1 , 890 cm ^-1 , and 950 cm ^-1 bands were observed, while for COD 610 cm ^-1 , and 920 cm ^-1.
Each band was observed. 780 cm ^-1 and 1320c
Each band of m ^-1 was observed in both COM and COD.
Moreover, the CaP 560 cm ^-1, 600 cm ^-1, and 1
Each band of 020 cm ^-1 was observed.

Among urinary tract stones, calcium oxalate (C
The combination of aOx) and calcium phosphate (CaP)
It is reported to be the most frequently observed and as much as 60 to 80% of the whole. It can be said that the infrared absorption spectrum of CaP contained in such a calculus has a great influence on the infrared absorption spectrum of urinary calculus CaUx, and particularly has a great influence in the range of 600 to 610 cm ^−1. You can Therefore, 600-6
If a band in the range of 10 cm ^-1 is used, COM and C
It will be appreciated that it is difficult to perform a quantitative analysis of COM and COD by analyzing common uroliths that normally contain CaP in addition to OD.

890 cm ^-1 , 920 cm ^-1 , and 950
infrared absorption in cm ^-1, the absorption is observed weaker than the infrared absorption at 780 cm ^-1, it is not suitable for this also accurate quantification. Therefore, a study was conducted focusing on the infrared absorption at 780 cm ⁻¹ that is not affected by CaP. The infrared absorption observed for the urinary calculus sample at 780 cm ^-1 is the absorption by COM and COD
It is considered to include absorption by. However, CO
It can be confirmed that M shows a sharp gradient in the range of 780 to 820 cm ^-1 , while COD has a gentle gradient in the range of 780 to 900 cm ^-1 . The difference in absorption between the pure COM sample and the pure COD sample changed from 780 cm ^{−1 as the} wave number increased, and the difference of 8 near 780 cm ^−1.
A maximum was observed at 00 cm ^-1 . The change in infrared absorption was small in COD and large in COM.

FIG. 2 shows the difference in absorption curve between calcium oxalate monohydrate and calcium oxalate dihydrate in the range of 700 to 900 cm ^-1 . COM
The absorption difference ab between the absorption spectrum of COD and the absorption spectrum of COD was examined for each wavelength. When the absorption difference ab was plotted for each wavelength, the results shown in FIG. 3 were obtained.
As shown in Fig. 3, COM and COD at 800 cm ^-1
It was found that the difference in absorption between and was the largest.

(3) Examination of Quantification Using Infrared Absorption Spectra 7 of each of pure COM standard sample, pure COD standard sample, and equal mixture of pure COM standard sample and pure COD standard sample
The infrared absorption spectrum of the absorption band at 80 cm ^-1 was analyzed. FIG. 4 is an infrared absorption spectrum of a pure COM standard sample,
FIG. 5 is an infrared absorption spectrum of the pure COD standard sample, and FIG. 6 is an infrared absorption spectrum of an equal mixture of the pure COM standard sample and the pure COD standard sample. First, in FIG. 6, a line connecting the valleys O and Q on both sides of the peak (P) is defined as a base line c. 780 cm on this line c
Two points at ^-1 and 800 cm ^-1 are designated as C and C ', respectively.

Draw a line CP between the C at 780 cm ^-1 and the absorption peak P, and further correspond to the absorption peak from the C'absorption parallel to the line CP and at 800 cm ^-1 . Draw a line C'-P 'to The distance from the C to the P is indicated by | C−P |. Where | C-P
| Is equal to | C'-P '|. Next, the point where the line C′-P ′ at 800 cm ⁻¹ intersects with the absorption spectrum is defined as R ′. Then, a line R'-R parallel to the line c (C'-C) is drawn. R is the point on line CP at 780 cm ^-1 . Focusing on an intersection R'with the spectrum on the line (C'-P '), R "corresponding to R'is obtained in the same manner in FIG. Further, in FIG. 5, R ′ ″ corresponding to R ′ is similarly obtained. R ''
Is located near C'and is only a short distance from C ', while R'"is located near P ',
It is only a short distance from P '. That is,
The position of R'is affected by the ratio of COM / COD concentrations.

If the COM / COD concentration ratio increases, the position of R'will be closer to R "or C '. As the COM / COD concentration ratio decreases, the position of R ′ comes closer to R ′ ″ or P ′. The position of R ″ and the position of R ′ ″ are
It is affected by the purity and / or weight or KBr of the standards used in each measurement. ｜ C'-P '
| Is the sum of absorption of COM and COD ([COM + CO
D]), | C'-R '|
COD absorption ([COD]), and | P'-R '|
Is the absorption of COM ([COM]) (note that the lines (C′-C), (R′-R) and (P ′ are
Since -P) is parallel, | C'-R '| is equal to | C-R | and | P'-R' | is equal to | P-R | (Figs. 4, 5 and 6). )), That is, for absorption, [COM + COD] = [COM] + [COD], then [COM] = [COM + COD] − [COD], and | P′−R ′ | = [COM] = | P− R | The absorption ratio COM / COD (Y) is Y = | P'-R '| / | C'-R' | = | P-R | / |
It can be seen that it can be represented by C−R |.

If the ratio of absorption of COM / COD is proportional to the ratio of concentration of COM / COD as predicted by Lambert-Beer's law, both are in a linear relationship. Therefore, the relationship between the measured infrared absorption value and the concentration ratio was investigated.
The absorption band at 800 cm ⁻¹ was used to determine the COD absorption ([COD]) value. The absorption band at 800 cm ⁻¹ was easy to read. The peak of absorption at 780 cm ^-1 was used as the value of the total absorption of COM and COD ([COM + COD]), and the ratio of the concentration of COM and COD was expressed as a percentage, and the measured values were plotted (FIG. 7). . From FIG. 7, assuming that the concentration ratio COM / COD is X, the following cubic equation relationship exists between the infrared absorption ratio of COM / COD and the concentration ratio of COM / COD when X> 0. Approved to be established. ^{Y = 0.163X 2 -1.484X Z + 4.778X} +
0.145

However, when 0 <X <1, the results shown in Table 1 were obtained. This is plotted in FIG. Then, the linear relationship is the infrared absorption ratio of COM / COD and COM / COD.
It was observed between the ratio of the concentrations of D. If this is shown by the formula,
The linear relationship is as follows: Y = 3.376X + 0.350 Then, the infrared absorption ratio Y'of COD / COM and COD / C
The results shown in Table 2 were obtained in the case of 0 <X '<1 between the OM concentration ratio X'. This is plotted in FIG. Then, a linear relationship was similarly recognized between the ratio of infrared absorption of COD / COM and the ratio of concentration of COD / COM. This can be expressed in the form of the following linear relationship: Y ′ = 0.143X ′ + 0.129

[0046]

[Table 1]

[0047]

[Table 2]

Further, when the experiment was repeated in the same manner as above, the following linear relationship was obtained when 0 <X <1. Y = 3.033X + 0.863 In consideration of the above result, when 0 <X <1, the following linear relational expression: Y = aX + b a = 2.720 to 3.689 b = 0 to 1.33
It is determined to be 0. Further, in the case of 0 <X <1, the experiment was repeated separately and the following linear relationship was obtained. Y '= 0.143 X' + 0.129 and Y '= 0.15
7X ′ + 0.104 As a result, when 0 <X ′ <1, the following linear relational expression: Y ′ = a′X ′ + b ′ a ′ = 0.130 to 0.170 b ′ = 0.080
It is determined that it becomes ~ 0.152. (4) Examination of Detection Sensitivity in Infrared Spectroscopy Using a pure COM standard sample and a pure COD standard sample, a sample containing 0% COM and 100% COD, that is, X = 0
Infrared absorption ratio Y = | P'-R '| / | C'-R' | = | P-R | / |
C−R | was repeatedly measured 10 times and examined. The results are shown in Table 3. Values are expressed as mean ± standard deviation (SD). Mean of confidence limits ± 25D. Was found to be 0.3899. Similarly, 100% COM and 0
% COD, i.e., the ratio of infrared absorption in the sample with X '= 0 Y' = | C'-R '| / | P'-R' | = | C-R | /
| PR | was repeatedly measured 10 times and examined. The results are shown in Table 4. The confidence limit was found to be 0.1355 (Table 4).

[0049]

[Table 3]

[0050]

[Table 4]

(5) Measurement of calculus sample in infrared spectroscopic analysis Three kinds of samples, ie, a sample consisting of one having COD <COM (sample 1), a sample consisting of one having COD ≠ COM (sample 2) and COD> A sample (Sample 3) made of COM was collected and uniformly used as a powder sample in the same manner as in (1). 1 for each sample
Infrared absorption spectrum is measured by repeating the same procedure 0 times,
The content rate of COM was calculated. The calibration curve was prepared in the same manner using a pure COM standard sample and a pure COD standard sample.
Measurements were made for both random and systematic errors. The results are shown in Tables 5-7 (accidental error) and Tables 8-10.
(Systematic error).

[0052]

[Table 5]

[0053]

[Table 6]

[0054]

[Table 7]

[0055]

[Table 8]

[0056]

[Table 9]

[0057]

[Table 10]

The effectiveness of the method of the present invention was confirmed. Each deviation was less than 5%.

(6) Effect of amount of calculus sample used on measurement Approximately 1 mg, 2 mg, 3 mg
About 4 mg and about 5 mg were weighed and about 240 potassium bromide powder (KBr for spectroscopic analysis, manufactured by Merck)
It was mixed with mg and formed into tablets (pellets). Infrared absorption spectra were measured in the same manner as in (1) above to determine the content rate of COM. The results are shown in Tables 11 and 12.

[0060]

[Table 11]

[0061]

[Table 12]

It is judged that the amount of the calculus sample used does not substantially affect the measurement result. However, in consideration of handling and the like, it is preferable to use a calculus sample of about 2 to 3 mg. Further, various methods such as pulverization can be applied as long as they do not adversely affect the measurement sample.

(7) Comparison of X-ray Diffraction Method and Method of the Present Invention for Quantifying Stone Samples Using the same stone sample, a Fourier transform infrared spectrometer (FT) was used.
-IR) (Model VALOR-III, manufactured by JASCO Corporation) was used to measure an infrared spectrum in the same manner as in (1) and (2) above, where the concentration ratio COM / COD was X and the absorption ratio was When COM / COD is Y and 0 <X <1, the following linear relational expression: Y = 3.376X + 0.350 is used, the infrared absorption ratio of COD / COM is Y ', and C
When the ratio of the OD / COM concentrations is X ′ and 0 <X ′ <1, the content ratio of COM was calculated using the following linear relational expression: Y ′ = 0.143X ′ + 0.129. For X-ray diffraction method, X
A line diffractometer XD-D1 type (manufactured by Shimadzu Corporation) was used. In the case of the X-ray diffraction method, data could not be obtained unless the sample was prepared carefully and uniformly, and the analysis took time.

[0064]

[Table 13]

From Table 13, it was confirmed that the method of the present invention has a very good correlation with the method by the X-ray diffraction method.

[0066]

INDUSTRIAL APPLICABILITY According to the present invention, quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopy can be performed by a simple method.
In the present invention, the quantitative determination of calcium oxalate / monohydrate and calcium oxalate / dihydrate is carried out quickly and easily by utilizing the infrared absorption spectrum at substantially 800 cm ^-1 .
And it can be done accurately. In particular, it is clinically useful because it can accurately quantify calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi over a wide range of content ratios. According to the invention, substantially 7
Infrared absorption spectrum at 80 cm ^-1 and substantially 8
Using the infrared absorption spectrum at 00 cm ^−1, the linear function between the amount of calcium oxalate monohydrate and the amount of calcium oxalate dihydrate in the stone and the infrared absorption was followed by a linear function that was easy to analyze. Relationships can be established and quantified.

[Brief description of drawings]

FIG. 1 shows an infrared spectrum by transmittance. A shows a pure calcium oxalate monohydrate standard sample, B shows a pure calcium oxalate dihydrate standard sample, and C shows an infrared transmission spectrum of calcium phosphate.

FIG. 2 shows an infrared absorption spectrum showing a difference in absorption characteristics between a pure calcium oxalate / monohydrate standard sample and a pure calcium oxalate / dihydrate standard sample.

FIG. 3 shows the relationship between the value of ab in FIG. 2 and the wave number of the infrared absorption spectrum.

FIG. 4 shows an infrared absorption spectrum of a pure calcium oxalate monohydrate standard sample.

FIG. 5 shows an infrared absorption spectrum of a pure calcium oxalate dihydrate standard sample.

FIG. 6 shows an infrared absorption spectrum of a sample of an equal mixture of a pure calcium oxalate / monohydrate standard sample and a pure calcium oxalate / dihydrate standard sample.

FIG. 7 shows the relationship between the COM / COD concentration ratio and the COM / COD infrared absorption ratio.

FIG. 8 is C when the concentration of COM / COD is less than 1.
1 shows the relationship between the OM / COD concentration ratio and the COM / COD infrared absorption ratio.

FIG. 9 shows C when the COD / COM concentration is less than 1.
1 shows the relationship between the ratio of OD / COM concentration and the ratio of COD / COM infrared absorption.

Claims (7)

(57) [Claims] 1. In the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in a living stone by infrared spectroscopic analysis, the peak of infrared absorption spectrum substantially appearing at 780 cm ^-1 and Using the infrared absorption spectrum up to 900 cm ⁻¹ on the high wave number side near the absorption peak, the amount of calcium oxalate / monohydrate and calcium oxalate / dihydrate and the infrared absorption The relationship between the spectrum and the first-order equation (linear equation) is obtained, and based on this relationship, calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones are quantified. how to. 2. In the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in a living stone by infrared spectroscopy, the peak of infrared absorption spectrum substantially appearing at 780 cm ^-1 is shown. A baseline is set that connects the bottoms of the spectral valleys on both sides of the sandwich, and the peak and the absorption peak, and the high wavenumber side near the absorption peak and up to 900 cm ^-1 Using the band of the infrared absorption spectrum in, the ratio of the concentration of calcium oxalate / monohydrate to calcium oxalate / dihydrate and the calcium oxalate / monohydrate and oxalic acid based on the baseline The relationship between the infrared absorption ratio of calcium dihydrate and the infrared absorption ratio was calculated according to a linear equation (linear equation), and based on this relationship, calcium oxalate monohydrate and oxalic acid oxalate in living stones were found. The method according to claim 1, wherein the amount of lucium dihydrate is determined. 3. The wave number near the absorption peak is substantially
The method according to claim ^1, which is 800 cm ^-1 . 4. Infrared absorption spectrum appearing at 780 cm ^-1 in the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in living stones by infrared spectroscopy.
Based on this relationship, the content ratio of calcium oxalate / monohydrate and calcium oxalate / dihydrate was measured by using the peak of Koutor and the absorption band at substantially 800 cm ^-1 . A method comprising quantifying calcium oxalate / monohydrate and calcium oxalate / dihydrate in urinary calculi. 5. In the quantitative analysis of calcium oxalate / monohydrate and calcium oxalate / dihydrate in a living stone by infrared spectroscopy, the peak of infrared absorption spectrum substantially appearing at 780 cm ^-1 and Utilizing the infrared absorption spectrum substantially appearing at 800 cm ^-1 , the 780 cm ^-1 based on the baseline of the peak of the infrared absorption spectrum appearing at 780 cm ^-1
Assuming a parallel homology between cm ^-1 and 800 cm ^-1 ,
^-1 shows the infrared absorption spectrum of calcium oxalate
The infrared absorption spectrum which is assumed to be the amount of infrared absorption of dihydrate and appears at 780 cm ^-1 is the sum of infrared absorptions of calcium oxalate monohydrate and calcium oxalate dihydrate in the sample. Between the content ratio of calcium oxalate monohydrate and calcium oxalate dihydrate and the measured infrared absorption spectrum appearing at 780 cm ^-1 of the linear equation (linear equation)
Calculating the relationship according to, and based on this relationship calcium oxalate monohydrate and calcium oxalate in living stones
The method according to claim 4, wherein quantification of dihydrate is performed. 6. Calcium oxalate monohydrate (COM) and calcium oxalate in urinary calculi by infrared spectroscopy
Substantially 800 cm ^-1 in quantitative analysis of dihydrate (COD)
The absorption band at is the infrared absorption of COD and is substantially 7
The absorption band at 80 cm ^-1 is the infrared absorption of the sum of COM and COD. (1) The concentration ratio COM / COD is X, the absorption ratio COM / COD is Y, and when 0 <X <1, The following linear relational expression: Y = aX + ba = 2.720 to 3.689 b = 0 to 1.330 (2) The infrared absorption ratio of COD / COM is Y ', the ratio of COD / COM concentrations is X', If 0 <X '<1, use the following linear relational expression: Y' = a'X '+ b' a '= 0.130 to 0.170 b' = 0.080 to 0.152 and use oxalic acid in urinary tract stones. The method according to claim 5, wherein the quantitative analysis of calcium / monohydrate and calcium oxalate / dihydrate is performed. 7. The method of claim 6 wherein a = 3.376, b = 0.350, a '= 0.143 and b' = 0.129.