УДК 636.028:574.24:591.044:591.111.1
PHYSIOLOGICAL RESISTANCE OF ERYTHROCYTES TO ETHANOL STRESS ON THE BASIS OF APPLYING PREPARATIONS FROM ROE ANTLERS
A.D. Korshunov, N.F. Ivankina
The effect of preparations from roe antlers on membranes of peripheral blood erythrocytes was studied in the conditions of ethanol stress in females of nonlinear white rats. The membrane-protective properties of biologically active substances from roe antlers were revealed. The injections by alcoholic extract from antlers powder were more effective in comparison with the addition this preparation to feed.
Key words: antler, stress, hemolysis, erythrocytes, resistance.
Preparations of non-ossified horns (antlers) of the Northern Forest deer (Rangifer tarandus) and the spotted deer (Cervus nippon) are widely used in traditional oriental and European medicine (1-4). These and other ungulates - elk, red deer, roe deer - inhabit in the Far East. The biological activity of preparations from elk antlers is confirmed by literature data (5), but we haven’t found any data on the chemical composition and biological activity of roe antlers.
Acid resistance of erythrocytes can be used as a stress marker for its close correlation with standard indicators of stress reaction, and its determination is technically simple (6).
The purpose of this work was studying the influence of preparations from roe antlers on membranes of peripheral blood erythrocytes in the conditions of ethanol stress.
Methods. The object of the study were the females of nonlinear white rats weighing 180-200g each one. In this work, we used the fine powder and alcoholic extract of roe antlers, obtained from the hunting areas of the Amur Region. The powder was extracted with 50% ethanol at a ratio of 1:1. Immediately before injection, the extract was diluted with water to a concentration of 35%, thus avoiding protein denaturation of tissues at parenteral introduction of the preparation.
The animals were divided into 4 groups of 6 rats in each group. Rats of the I st (control), II and IV groups were fed a standard diet (the Ministry of Health order № 163 from 10.03.1966) during 30 days. Group III: the powder from roe antlers (30 mg per 100 g of body weight) was added to the feed every second day. Group IV: the alcoholic extract of roe antlers powder (0.1 cm3 of 35%) was subcutaneously injected on the 29 th and 30 th days. On the 30 th day, 1 cm3 of 96% ethanol was introduced intragastrally to the rats of groups II-IV, and 1 h later all the animals were slaughtered by decapitation. Blood was collected in heparinized tubes.
The effects of the preparation on cell membranes were estimated from the resistance of erythrocytes, which was determined by the modified method of acid erythrograms (7). 0,004 N hydrochloric acid in saline solution was used as hemolytic. Blood (50 ml) was mixed with saline solution (50 cm3), obtaining the suspension of erythrocytes in a dilution of 1:1000. Investigations were carried out at room temperature.
The optical density of suspension of erythrocytes was measured colorimetrically (fotoelectrocolorimeter KFK-2 by "Zagorskij Optical and Mechanical Plant", Russia) with a red light filter and cuvettes with optical path length of 10 mm. The suspension of erythrocytes from a single sample was placed in a cuvette, and HCl (2 cm3 0,004 N) was added. Time elapsed for complete hemolysis of the suspension was taken as a measure of resistance of erythrocytes.
The results were processed statistically using MS Excel and STAT-1software packages.
Results. In group I (control), the time of hemolysis of erythrocytes was 16,08 ± 1,20 min, in group II it was significantly less - 14,00 ± 1,14 min (p <0,05). This decrease in resistance of erythrocytes in conditions of ethanol stress is consistent with the results obtained by other authors (8-10). The ethanol transport occurs by concentration gradient via cell membrane channels, and to a less extent - through dissolution in lipid layer. Being dissolved in water and - partly - in lipids of cell membrane and subcellular structures, ethanol causes their fluidization (increase of membrane fluidity) (10). Ethanol also affects conformation of protein molecules, thereby violating their ability to proper operation (11). The negative effects of ethanol on membranes is enhanced by its main metabolite - acetaldehyde, whose covalent interaction with amino- and sulfhydryl- groups of proteins of blood cells result in destruction of their structural organization and functional activity (11).
In groups III and IV, time of hemolysis of erythrocytes was 17,33 ± 0,82 and 22,66 ± 1,17 min, respectively. These values were significantly higher than in group II (p <0,001). Feeding of the roe antlers powder (group III) on the background of intragastric introduction of ethanol almost didn’t affect the time of hemolysis of erythrocytes (p> 0,05).
Thus, the biologically active substances from roe antlers perform membrane-protective properties in conditions of ethanol stress; injections of the alcoholic extract of powdered antlers are more effective than oral introduction of antlers powder.
REFERENCES
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10. Prokop’yeva V.D., Bokhan N.A. and Kondakova I.V., The Effects of Ethanol and Acetaldehyde on Microviscosity of Erythrocyte Membranes in Patients with Alcoholism at Various Treatment Stages, Bull. eksp. biol. i med., 2000, vol. 129(1), pp. 90-92.
11. Goldstein D.B., Pharmacology of Alcohol, New York, 1983.
Far Eastern State Agricultural University, Blagoveschensk 675005, Amur reg., Russia |
Received June 18, 2008
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