JPH01174940A - Measurement of hemoglobin - Google Patents

Abstract

PURPOSE:To directly and continuously measure the variation in the amount of hemoglobin in blood of the principal tissue of a living body from the living body, by measuring the change in the absorbancies of two wavelengths selected in a wavelength region where only the absorbancy change caused by hemoglobin is generated. CONSTITUTION:In a wavelength region where only the change in absorbancy caused by hemoglobin is substantially generated, two specific different wavelengths lambda1, lambda2 are selected and lights having these wavelengths are allowed to directly irradiate the tissue of a living body to measure absorbancy changes DELTAA1, DELTAA2 with respect to the respective wavelengths. On the basis of these absorbancy changes DELTAA1, DELTAA2, absorptivity coefficient k1 preliminarily obtained by a specific wavelength, the absorptivity coefficient k1' of oxygenation type hemoglobin at a wavelength lambda1, the absorptivity coefficient k2 of deoxygenation type hemoglobin at the wavelength lambda1, the absorptivity coefficient k2' of oxygenation type hemoglobin at a wavelength lambda2, the absorptivity coefficient of deoxygenation type hemoglobin at the wavelength lambda2 and absorptivity coefficient k3 at the isosbestic point (wavelength lambda3) of oxygenation and deoxygenation hemoglobins, the variation in the amounts of oxygenation and deoxygenation type hemoglobins and/or the variation in the amount of total hemoglobin is measured.

Description

DETAILED DESCRIPTION OF THE INVENTION (6) Industrial Application Field This invention relates to a method for measuring hemoglobin. More specifically, the present invention relates to a method for directly measuring changes in the amount of hemoglyceride in the blood of a living body using light of a specific wavelength in the near-infrared region.

(B) Conventional technology Conventional methods for measuring hemoglobin (hereinafter abbreviated as Hb) in blood include a method in which blood is taken out of the living body and measured, and a method in which it is measured without taking blood out of the living body. There is. For the latter method.

There are two methods: percutaneous measurement by heating the skin to create arterial blood and attaching an oxygen electrode to the area; optical methods using oximeters such as earpiece oximeters and pulse oximeters; and methods using reflected light. The oxygenated (oxy)-deoxygenated (deoxy) spectrum of Hb, u
There are known methods for measuring fluctuations in the amount of hemoglobin based on changes in hemoglobin.

(c) Problems to be solved by the invention However, the percutaneous measurement method described above cannot be used at the same location for a long time, and the information obtained can only be obtained from peripheral blood vessels. It is not possible to know the state of blood in major tissues such as the body.

On the other hand, in the method of using an oximeter, pulse oximeters are widely used, but they can only measure blood in the arteries at the fingertips,9 and cannot directly measure blood in the main tissues of the body such as the head and internal organs. cannot be measured.

Furthermore, in the method based on spec)/l/ change, the absorbance change due to blood volume fluctuation overlaps with the absorbance change due to oxy-deoxy.

In addition, when measuring major tissues, especially the head, cytochrome a
The spectral fluctuations associated with changes in the redox state of a3 (cytaas) overlap;
Since the optical path and optical path length differ depending on the wavelength, distortion occurs in the spectrum, which poses problems such as the inability to easily measure parameters corresponding to the amount of hemoglobin.

This invention was made in view of the above circumstances.92 Hemoglobin measurement that enables direct and continuous measurement of hemoglobin level fluctuations in blood in the main tissues of a living body such as the head and internal organs using an optical method. It seeks to provide law.

B) Means for Solving the Problems Thus, according to the present invention, two different specific wavelengths λ1 and λ2 are selected in the wavelength range where only the absorbance change due to hemoglobin occurs, and light of these wavelengths is applied to living tissue. Absorbance change △A for each wavelength by direct irradiation
1 and ΔA2, and the absorbance changes ΔA1 and ΔA2 and the extinction coefficient 5 previously obtained at the above-mentioned specific wavelength. kl: Extinction coefficient of oxygenated hemoglobin at wavelength λl.

k'l: Wavelength λ! Extinction coefficient of deoxygenated hemoglobin in

k22 Extinction coefficient of oxygenated hemoglobin at wavelength λ2.

k'22 Extinction coefficient of deoxygenated hemoglobin at wavelength λ2.

and the extinction coefficient of 3 at the isosbestic point (wavelength λ3) of oxygenated hemoglobin and deoxygenated hemoglobin, the variation in the amount of oxygenated hemoglobin in the irradiation optical path, the variation in the amount of deoxygenated hemoglobin, and/or Alternatively, a hemoglobin measuring method is provided, which is characterized by measuring fluctuations in total hemoglobin amount.

This invention is an optical method that uses wavelengths in the near-infrared region for biological tissues, by selecting two specific different wavelengths (λl, λ2) from a wavelength region of 700 nm or more that cause only a change in absorbance due to hemoglobin. By using it as The present invention is characterized in that it is a method capable of non-invasively monitoring the tissue by directly irradiating the tissue with light of the above wavelength.

The two different specific wavelengths used in the method of the present invention are selected from a wavelength range of 7QQnm or more in which substantially only a change in absorbance due to hemoglobin occurs, and the wavelength range is particularly preferably 700 to 780 nm. Regarding this, reference will be made to the description of the embodiment described later. In addition, even outside the above-mentioned preferred wavelength range, the concentration of inhaled oxygen in the main tissue of the living body to be measured is extremely small, so long as the SBEK)/7 variation accompanying the redox change of cytochrome aa3 cannot be ignored.

Since the spectrum)/L/ variation can be ignored compared to the spectrum variation accompanying oxygenation-deoxygenation of hemoglobin, the method of the present invention can be fully applied.

In the method of this invention, two wavelengths are selected from the above wavelength range.
The wavelength is selected from a combination that provides a large difference in absorbance and less wavelength dependence due to scattering, etc.1, for example, 700 nm.
Combinations of 730nm, 75Qnm and 713Qnm, etc. are preferred, but are not limited to these.

Further, the point at which the absorbance of oxygenated hemoglobin and deoxygenated hemoglobin become the same is around the wavelength of 8051 m.

In the method of this invention, the change in absorbance is measured.

Spectrophotometers and detectors commonly used in this field can be used as they are. However, in the measurement optical path from the light source to the detector, a measurement section configured to directly irradiate the specific wavelength light selected as described above to the biological part to be measured is set.

The measurement optical path and measurement section as described above can be constructed using an optical fiber or the like.

The method of the present invention is used within a physiological range in which the Lambe-Az)-Beer law holds between fluctuations in the amount of hemoglobin and changes in absorbance. That is, the irradiation optical path (optical path length: d) with specific wavelengths (λl, λ2, λ3) to the biological tissue
Changes in each amount to be determined in: Oxygenated hemoglobin (HbO
z) Volume fluctuation, deoxygenated hemoglobin (Hb) volume fluctuation, and total hemoglobin (IHb) volume fluctuation are respectively △
(HbOz), △(Hb), △(IF(b:)), and the extinction coefficient of the tissue by the irradiation light of the above specific wavelength, that is, the extinction coefficient of oxygenated hemoglobin at wavelength λ1: k, , also at wavelength λ1 The extinction coefficient of deoxygenated hemoglobin at wavelength λ2 is 2:2.
Extinction coefficient of deoxygenated hemoglobin in 2: ni',
If the extinction coefficient of navel globin at the point (wavelength λ3) where the absorbance of oxygenated hemoglobin and deoxygenated hemoglobin are the same (wavelength λ3) is 2:3, then each wavelength λ1. λ2 or λ
Amount of change in absorbance over time due to 3 ΔA1 + ΔA2 and ΔA
3 is △A + = k+△(HbOz:] +kt'△(H
b) −■△Ax = kz△(H
bOz:)+kz'△CHb) -■
△A3shikj△(HbOz) + ksΔ[:Hb)=
is within the physiological range where a linear relationship expressed by 3Δ[II(b) -■ holds true.

According to the above formulas ■ and ■, each quantity fluctuation is △[H'bO2] = (k2'△A+ - +7 Hu)/
(ki k2'-kzk1') -■△CH
b) = (k2△Ax- to 1△A2)/(kzk1'-k
tk;) −■△(IF(b) = ((k
z'-kz)mu1-(k+'-kl)△nn (klk2
'-2kl') -■, and by using 3, the following equation is calculated.

ζ where kl'/3 e k2'/3 is the ratio of extinction coefficients of deoxygenated hemoglobin at λ1 and λ2 and λ3.

This ratio can be determined by, for example, the ratio of the change in absorbance of a rat skeletal muscle that has been completely anaerobic. In other words, under completely anaerobic conditions in rats (10% 02 inhalation), blood with different hematocrit (hereinafter abbreviated as Hct) was flowing, and λ was adjusted each time.
1. λ2. Plot the absorbance obtained by alternately irradiating light with a wavelength of λ3, and calculate λl, λ2 and λ3 based on )lct.
Create a correlation graph of absorbance. The coefficient ratio can be measured by finding the ratio of absorbance changes in the linear component between wavelengths.

Also, kl/3. kg/ks can also be determined by conducting a similar experiment under completely aerobic conditions.

All of the above coefficients can also be calculated in an in vivo system.

If a certain state is n, a completely aerobic state (arteriovenous blood oxygen saturation of 100) is a, and a completely anaerobic state (arteriovenous blood oxygen saturation is O) is (HbOz)n.

(Hb)n, (IHb:ln is expressed as follows.

(HbOz)J3=△(HbOz)*-bKi= ((
k;/ks)ΔA1n-'(kt'/IC5)ΔA
zn-b)/K"((k2'/to3)lOgI+/It
(kt/)C3)A! ogIz/Izn]/K(H
b, lJ<3=Δ(Hb)vt<<.

=(-(kz/ki)△A+n-a+ (3 to k1/)
△Az −”)/ni−(−(kz/ICs)logII
a/ITo+(3MogI2she2° to k+/)/K[
et(b)hk3 = (HbOz)nkko+ (Hb
]nk3 where △(HbOz)n-b: Amount of change in oxygenated hemoglobin from state gb △(Hb)na-a: Amount of change in deoxygenated hemoglobin from state a ΔA1''-b: When in state A Absorbance change function I for wavelength λ1
n = amount of transmitted light with wavelength λl in state n Therefore, it is necessary to change from the original state to a completely aerobic state. The absolute amount of converted hemoglobin is determined.

(e) Examples Example 1 FIG. 5 shows a block diagram of an example of an apparatus for implementing the hemoglobin measurement method of the present invention. This device is a block diagram of a model experiment conducted using rats. In the figure (
1) is a flow control, (2) is a humidifier, (3)
is an artificial respirator, (4) is a light source, (5) is a monochromator, (6) is a photomultiplier tube (PM), (7) is a LOG converter, (8) is an analog/digital converter, (9) )HaC
PU, ([(l is high voltage (HV).

The above (1), (2), and (3) are mechanisms for adjusting the rat's body from an aerobic state to an anaerobic state.

(3) is therefore configured to be connected to the rat's trachea. Further, an optical path (a) is formed by an optical fiber from the light source (4) to the above PM (6) via the measuring section (A), and this optical path is interrupted at the above measuring section (4). An irradiation end and a light reception end are set. The body tissue to be measured is sandwiched between the irradiating end portion and the light receiving end portion so that the biological tissue to be measured can be directly measured. Although the monochromator (5) is located after the measurement section (A), it can also be used for front spectroscopy. Although a normal monochromator may be used, an interference filter may be used to select only the intended wavelength light and irradiate it alternately.

A case will be described in which the oxygen saturation of hemoglobin (hereinafter abbreviated as Hb) is measured using a rat head transmission spectrum obtained using the apparatus configured as described above. Generally, in the head, it is a biological material that has a characteristic absorption peak in the near-infrared region.

Hb and cytochrome aa3 (hereinafter abbreviated as cytaa3) are considered. Therefore, rats were placed in an aerobic state B (95%025
%CO2 inhalation) and its absorption rate) /L/(a)
Measure.

After that, in an anaerobic state (100% N2 inhalation), the absorption nupek) / I / (b) was measured in the same way, and regarding these results, the absorption nubek) / 1 / (a) was compared to the baseline (■)
The difference between the absorption spectra (subek)/v(■) and the absorption spectra (b) are shown in FIG. 6 (B). This result is quite different from the result of similarly measuring only Hb isolated from rats (Fig. 7).

Next, the rats were perfused with fluorocarbon (artificial blood) to replace Hb and become Hb-free.

Regarding this, the same baseline (■) and difference spectrum (■) as above were measured, and the results shown in Figure 6 (~) were obtained.
This clearly shows the fluctuations in the redox level of cytaa3 in the rat head.

Therefore, from 700 to 7801 m, it can be considered that there is almost no change in the spectrum due to changes in the redox level of cytaaB.

From the above, it can be said that the data obtained by setting the measurement wavelength in this wavelength range depends only on the oxygenation-deoxygenation of Hb.

Therefore, the 0 point of H'bO2 concentration was set for the measurement in rats under completely anaerobic conditions with 0% 02 inhalation, while 100%
Measured under fully aerobic conditions by inhalation of %02
It is important to set the Nuban point of bOz concentration.

It becomes possible to quantify the oxygen saturation of hemoglobin.

Example 2 Using the apparatus shown in Fig. 5, 700 nm for λl, 730 nm for λ2, and 805 nm for λ3') * Ra
Transmitted light photometry of the thigh skeletal muscle was performed.

First, in a Rat head perfusion experiment using the above two wavelengths, a completely aerobic state (100% 02 inhalation) and a completely anaerobic state (
The results of perfusing blood in the range of 15 to 50% HCl under conditions of 0% 02 inhalation] and measuring changes in absorbance are shown in the first
As shown in the figure. As shown in FIG. ).

Therefore, specific wavelengths: 700nm, 730nm, 80nm
Regarding the variation in oxygenated hemoglobin amount: △ (HbOz), the variation in deoxygenated hemoglobin amount: △ (Hb), and the variation in total hemoglobin amount: △ (IHb) in the irradiation optical path (optical path length d) at 5 nm, the above-mentioned The formulas ■～■ hold true.

Next, in the formula ■'~■', '/, , k2'/,
(700 nm and 73 nm for deoxygenated hemoglobin
The ratio of extinction coefficients at 0 nm and 805 nm) is determined by the ratio of changes in absorbance of completely anaerobic rat skeletal muscle.

This was done in rats under completely anaerobic conditions (0% 02 inhalation).
700 nm each time while flowing blood with different lct.

Plot the absorbance obtained by alternately irradiating wavelength light of 730 nm and 805 nm) based on let <700 nm
.

This is determined by creating a correlation graph between absorbance at 730 nm and 805 nm and measuring the slope of the resulting graph. FIG. 2(a) shows the above correlation graph. From this graph,
kl'/3=1.50 k2'/k3= 1.23
It became.

Furthermore, e kl/k3 m k2/k3 can also be determined by conducting a similar experiment under completely aerobic conditions (see Figure 2 (b)). From this graph, kl/
3=0.80, kt/k3=0.86.

Therefore, ■′, ■′, and ■′ are respectively as follows.

△(HbOz) k3= 4.23ΔA? 0O-5,1
5ΔA730 −■1Δ(Hb)ks = 2.92
△A700-2.75ΔA73G -■'△(IH
b]ks = 1.31△A700-2.41△A73
0 -■' again. The rat thigh was irradiated with light, and the amount of oxygenated hemoglobin CHbOz, l ks , the amount of deoxygenated hemoglobin ()Ib]ki, and the amount of total hemoglobin (
IF (b″lka and 802 absorbs 95% of inhaled gas 02
Figure 3 shows how it behaves when the CO2 is lowered from 5% CO2 to 0%02.

It can be seen that each quantity is well monitored by the optical parameters that we have calculated.

FIG. 4 shows the shift of the present invention.
This graph explains that it is possible to monitor changes in the amount of oxygenated hemoglobin and changes in the amount of deoxygenated hemoglobin due to fluctuations in the intake gas at the Ra1 head using the m wavelength bear. According to this figure, when switching from oxygen inhalation to nitrogen inhalation, △(Hbo2)ki decreases significantly, and Δ(
Hb) shows a significant increase in k3,
The fact that when the jugular vein is severed after being in a completely anaerobic state (O in 802), HbOz hardly decreases and only the skin significantly decreases means that at this time, hemoglobin is mostly in the reduced form. Match.

(f) Effects According to the present invention, changes in the amount of oxygenated hemoglobin, changes in the amount of deoxygenated hemoglobin, changes in total hemoglobin (i.e., blood volume), and changes over time in oxygen saturation in living tissues within the irradiation optical path are monitored. be able to.

[Brief explanation of the drawing]

Figure 1 shows fully aerobic conditions (a) and fully anaerobic conditions ('
Figure 2 is a diagram showing the relationship between ct and absorbance in Rat skeletal muscle at (b). Figure 2 is a diagram showing the correlation between absorbance changes between wavelengths in fully aerobic conditions (a) and fully anaerobic conditions (b). , inhalation 0
Figure 4 shows the change in the state of Rat femoral hemoglobin as the concentration decreases.
Figure 5 is a behavioral diagram of the amount of oxygenated hemoglobin and deoxygenated hemoglobin in one head. FIG. 6 is a diagram of an apparatus for carrying out the method of the present invention, and shows the transmission spectrum of a rat under head perfusion, and the difference spectrum between the absorption spectrum under aerobic conditions and the absorption spectrum under anaerobic conditions. The graphical diagram, Figure 7, is the Figure 6 phase diagram for rat liberated hemoglobin. Shaku tLt bone 祢荀'＞t=B keri Hctt's guan'i kezu n skin-! c rfJq sound exhibition 1 Thailand seasonal news 8i Wazu dormitory 7 Figure 7oo 3oo 9θ0 mark ゑ→

Claims (1)

[Scope of Claims] 1. In a wavelength range in which spectral fluctuations associated with changes in the redox state of cytochrome aa3 are negligible compared to spectral fluctuations associated with oxygenation-deoxygenation of hemoglobin, and substantially only absorbance changes due to hemoglobin occur. Select two different specific wavelengths λ_1 and λ_2, directly irradiate living tissue with these wavelengths, measure the absorbance changes ΔA_1 and ΔA_2 for each wavelength, and measure the absorbance changes ΔA_1 and ΔA_2 for each wavelength.
_1 and ΔA_2, and the extinction coefficient obtained in advance at the specific wavelength; k_1: extinction coefficient of oxygenated hemoglobin at wavelength λ_1, k'_1: extinction coefficient of deoxygenated hemoglobin at wavelength λ_1, k_2: extinction coefficient at wavelength λ_2 Extinction coefficient of oxygenated hemoglobin, k'_2: Based on the extinction coefficient of deoxygenated hemoglobin at wavelength λ_2, and the extinction coefficient k_3 at the isosbestic point (wavelength λ_3) of oxygenated hemoglobin and deoxygenated hemoglobin A method for measuring hemoglobin, which comprises measuring fluctuations in the amount of oxygenated hemoglobin, fluctuations in the amount of deoxygenated hemoglobin, and/or fluctuations in the amount of total hemoglobin in the irradiation optical path. 2. The wavelength range is 700 to 780, where the spectral fluctuations associated with changes in the redox state of cytochrome aa3 are negligible compared to the spectral fluctuations associated with oxygenation-deoxygenation of hemoglobin.
The hemoglobin measurement method according to claim 1, wherein the hemoglobin measurement method is nm.