Methemoglobin reductase activity of nuclear erythrocytes of the sea ruff (Scorpaena porcus, Linnaeus, 1758) under normal and oxidative stress conditions (in vitro experiments)

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Abstract

A method for evaluating the methemoglobin reductase activity of nuclear erythrocytes of teleost fish is proposed. The work was performed on a bottom marine species (Scorpaena porcus, Linnaeus, 1758). In an in vitro experiment, the erythrocyte suspension of this type was weighed in solutions with different concentrations of NaNO2, after washing from this compound, the process of methemoglobin (MetHb) reduction was studied for 150 min. The functional state of hemoglobin was judged by the results of spectral analysis. The study of the kinetics of MetHb reduction showed that the dependence was well described by the equation of an exponential function with a coefficient of determination (R2) greater than 0.9. The nature of the dependence remained at different levels of oxidative stress. This made it possible to calculate the velocity constant of the first order k (25 oC). In this species, it was 5.75–6.45 (10–3) min–1, which slightly exceeded the known values for mammals, and was close to the data obtained for other species of bony fish. It was found that the MetHb-reductase activity of the nuclear erythrocytes of the sea ruff increased with a rise in the oxidative load. At a concentration of 6.0 mM NaNO2, it was 1.73 ± 0.21 µM MetHb min–1 g–1 Hb. It is assumed that this is due to the process of deoxygenation of hemoglobin.

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About the authors

A. A. Soldatov

Federal Research Center “A.O. Kovalevsky Institute of Biology of the South Seas of the Russian Academy of Sciences”; Sevastopol State University

Author for correspondence.
Email: alekssoldatov@yandex.ru
Russian Federation, Nakhimov av., 2, Sevastopol, 299011; Universitetskaya str., 33, Sevastopol, 299053

N. T. Shalagina

Federal Research Center “A.O. Kovalevsky Institute of Biology of the South Seas of the Russian Academy of Sciences”

Email: alekssoldatov@yandex.ru
Russian Federation, Nakhimov av., 2, Sevastopol, 299011

V. N. Rychkova

Federal Research Center “A.O. Kovalevsky Institute of Biology of the South Seas of the Russian Academy of Sciences”

Email: alekssoldatov@yandex.ru
Russian Federation, Nakhimov av., 2, Sevastopol, 299011

T. A. Kukhareva

Federal Research Center “A.O. Kovalevsky Institute of Biology of the South Seas of the Russian Academy of Sciences”

Email: alekssoldatov@yandex.ru
Russian Federation, Nakhimov av., 2, Sevastopol, 299011

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Supplementary files

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2. Fig. 1. Spectral characteristics of black scorpionfish hemolysates under normal conditions and under oxidative stress (a – at 1.5 mM NaNO2; b – at 3.0 mM NaNO2; c – at 6.0 mM NaNO2; green spectrum – before adding NaNO2; red spectrum – immediately after adding NaNO2; blue spectrum – 150 min after washing away NaNO2)

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3. Fig. 2. An example of the kinetic curve of MetHb recovery after the removal of oxidative stress (NaNO2) (a – normal scale, b – logarithmic scale)

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4. Fig. 3. Kinetic characteristics of the MetHb reduction process after removal of oxidative stress of varying intensity (1 – at 1.5 mM NaNO2; 2 – at 3.0 mM NaNO2; 3 – at 6.0 mM NaNO2)

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5. Fig. 4. The reaction rate constant (k) of the first order (1 – at 1.5 mM NaNO2; 2 – at 3.0 mM NaNO2; 3 – at 6.0 mM NaNO2)

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6. Fig. 5. MetHb-reductase activity of nucleated erythrocytes of the black scorpionfish under oxidative stress of different intensity (1 – at 1.5 mM NaNO2; 2 – at 3.0 mM NaNO2; 3 – at 6.0 mM NaNO2)

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