RU2133033C1 - Method of diagnosis of organism pathology - Google Patents

Abstract

FIELD: biology, medicine. SUBSTANCE: method involves determination of reaction rate of superoxide anion-radicals with different biological fluids using semiconductive chemical sensor of n-type (zinc oxide). EFFECT: decreased time of study. 2 tbl, 2 dwg

Description

 The invention relates to biology and medicine and can be used both for the diagnosis of pathology and for the study of the vital processes of the body.

It is known that the superoxide anion radical (O ₂ ^- ) (COAP) is actively involved in the normal and pathological processes of the body. With his participation, enzymatic reactions catalyzed by oxyreductases, the interaction of hemes with oxygen, and problems of phagocytosis are associated [1].

 However, in the above work, COAP reactions with biological fluids obtained directly from the body were not studied, and the specificity of COAP reactions with components of biological fluids of a healthy and diseased organism was not revealed. Such studies have not been conducted because of the difficulty in determining the concentration of COAP and reaction products.

 The closest solution to the proposed invention is the work [2], where it is shown that the antioxidant potentials, defined as the reaction rate of red blood cells of healthy mice and cancer patients with organic hydroperoxide, differ. However, in [2], the antioxidant potential is introduced artificially by the reaction of red blood cells with an oxidizing agent that is not involved in the life of the body. This method was used due to the impossibility of physical methods to distinguish between oxidized red blood cells and non-oxidized, as well as the lack of rapid methods for controlling free radical products in the liquid phase.

 The technical task of this invention is the rapid determination of the direct value of the antioxidant potential of biological fluids and their components.

This is achieved by the fact that for the express determination of the direct value of the antioxidant potential of biological fluids and their components, a direct method is used to measure the relative reaction rate of these substances with COAP, i.e. an oxidizing agent directly involved in the life of the body. For express determination of the oxidation rate, a semiconductor chemical sensor is used, whose stationary conductivity is proportional to the concentration of COAP in the liquid medium. The stationary concentration of COAP in biological fluids is supported by the corresponding enzymes responsible for the formation and death of O ₂ ^- [1]. Consequently, the rate of interaction of COAP with biological fluids can characterize both viability and indicate pathological changes in it.

 The novelty of the present invention, in contrast to the prototype (the antioxidant potential is introduced artificially by the reaction of red blood cells with an oxidizing agent that is not involved in the life of the body) is the express determination of the direct value of the antioxidant potential of biological fluids and their components by direct measurement of the relative reaction rate of these substances COAP.

There are many works that speak of the important role of O ₂ ^- , which take an active part in normal and pathological processes of the body’s life, however, none of them investigated COAP reactions with biological fluids obtained directly from an animal organism. This allows us to conclude that the proposed invention meets the criterion of "inventive step".

As a COAP sensor, a semiconductor chemical sensor was used - a thin polycrystalline film of zinc oxide, which has proven itself in physical and chemical studies [3]. It turned out that in this case, as for other active particles, the relation between the concentration of COAP in the solution and the conductivity of the sensor
Δσ / σ ₀ = K [O $\genfrac{}{}{0ex}{}{\text{-}}{\text{2}}$ ] ^α ,
where Δσ = | σ-σ ₀ |, σ ₀ is the initial value of the sensor conductivity at zero COAP concentration σ is the stationary value of the sensor conductivity at a certain COAP concentration. The K-constant is determined experimentally for each sensor, α varies in the range 0 <α <1 and depends on the particular adsorbent-absorbate pair.

The COAP generation facility is an electrolyzer with a mercury cathode and a platinum anode, oxygen is bubbled through the cathode region. A solution of tetrabutyl ammonium perchlorate (PCTBA) in dimethylformamide (DMF) was used as an electrolyte. In such a system (PCTBA / DMF) it is known [4] that only COAP is formed in the cathode region, and its amount is related to the magnitude and duration of electrolysis by the Faraday law. The enrichment of the cathode region of COAP was performed for 2-6 hours at a current strength of 0.5 mA. The concentration of COAP was ≈ 10 ¹⁹ -5 • 10 ¹⁹ cm ^-3 .

 The invention is illustrated by the following examples.

 1. After electrolysis, a sample of 1-8 ml of a solution enriched in COAP was taken and transferred to a test tube, where the response of the conductivity of the sensor was determined.

 In FIG. Figure 1 shows the dependence of the relative electrical conductivity of the sensor on the concentration of COAP in a solution of PCTBA / DMF. From the calibration curve (Fig. 1) it can be seen that the sensor is a linear sensor of the concentration of COAP in a solution of PCTBA / DMF in the specified range of COAP concentrations.

 In FIG. Figure 2 shows the time it takes to establish the stationary electrical conductivity of the sensor with a stepwise change in the concentration of COAP in the PCTBA / DMF solution, which is 90 sec, which is convenient for measuring the reaction rate of COAP with various substances.

The lifetime (τ) of COAP in such a solution at a temperature of 20 ^o C is ≈ 6.3 hours, which is consistent with [1].

 2. The same as in paragraph 1, but to the selected sample enriched in COAP, add biological fluids in an amount (experimentally selected for convenience) of 0.4 vol.% And measure the relative reaction rate of COAP with blood, its components, urine or in other words determine the antioxidant potential (AOP) of the blood, its components, urine.

 Table 1 presents AOP of biological fluids. AOP values were normalized by AOP of blood of a healthy person (see the end of the description).

 Table 1 shows that the biological fluids studied by us have different AOP values. However, the experiment showed that the AOP of the blood of two practically healthy people have close values, but the cancer of the patient with AOP is 30% higher than that of a healthy one, which is shown in Table 1.

 Table 2 presents AOP of the urine of healthy and sick people. AOP normalization was carried out according to AOP of a practically healthy person (see the end of the description).

 As in studies with blood, AOP of the urine of two almost healthy urines is close.

 From the above data it follows that AOP can characterize both the state of a healthy organism and indicate a pathology in it.

Claims (1)

 A method for diagnosing an organism’s pathology by measuring the antioxidant potential of a biological fluid to increase the rate of its reaction with a chemically active substance, characterized in that for the expressive determination of the direct value of the antioxidant potential, the relative reaction rate of the superoxide anion radical with biological fluids is measured using a p-type semiconductor chemical sensor (zinc oxide).

Images