Therapeutic agent for preventing and treating chronic liver diseases

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to a therapeutic agent for preventing and treating chronic liver diseases. The above agent represents amaranth oil prepared by cold pressing of amaranth seed kernels and coats, to be used in a dose of 62.5 - 250 mg/kg of body weight.

EFFECT: declared invention provides higher positive effect on biochemical processes in blood and liver tissues in treating toxic hepatitis.

2 tbl

 

The invention relates to the field of pharmacology, namely, hepato-protective drugs and can be used in the development of new drugs of plant origin for the correction of cytolytic syndrome in the prevention and treatment of chronic liver diseases.

Known hepatoprotective drug Essentiale production "Rhone-Poulenc Rorer, USA/France; "Natterman, Germany, which consists of essential phospholipids (EFL) of high degree cleaning - diglyceride esters holinesterzoy acid and unsaturated fatty acid extract from soy beans. In addition, the table contains Riboflavin, thiamine, pyridoxine, cyanocobalamin, vitamin e, nicotinamide, Pantothenic acid (Sports Wiki.ru).

Known drug inhibits the processes of free radical oxidation of lipids, influence of physico-chemical properties of the membrane, reduces the viscosity of the lipid layer that modulates the activity of membrane-associated enzymes. Restores the function of cell membranes. Helps improve health of patients.

Known use as a prophylactic agent in toxic hepatitis seed oil, amaranth, obtained by extraction method at a dose of 0.5 ml per kg of body weight (Twin G.N., Turkish M.I., Polyakova-Semenova N and other Role processes freedoms is radikalnogo oxidation mechanism of hepatoprotective activity of oils from seeds of amaranth // Biomedicine. 2006. No. 2. S-112). It is established that the oil has a pronounced hepatoprotective effect, however, doses for the treatment of liver diseases is not installed.

In addition, the extraction method of obtaining oil from seeds of amaranth does not guarantee chemical purity, it contains phospholipids, Tocopherols, unsaturated fatty acids, squalene, phytosterols, esters of sterols, and reduces the effectiveness of therapeutic effects.

The objective of the invention is the expansion of the range of herbal medicines and the creation of a domestic product for the prevention and treatment of chronic liver diseases.

Technical result - increasing positive effect on biochemical processes in the blood and liver tissue in the treatment of toxic hepatitis.

The technical result is achieved that therapeutic agent for the prophylaxis and treatment of chronic liver diseases, representing amaranth oil, the latter obtained by cold pressing of the embryo and membranes seeds of amaranth, which is used at a dose of 62.5-250 mg/kg of body weight.

Amaranth oil, obtained by cold pressing contains phospholipids, Tocopherols, unsaturated fatty acids, squalene, phyto-sterols and esters of sterols in the following ratio, wt.%: phospholipids 7-9, tocopherol 0,3, unsaturated LM is nye acid 51,25, squalene 3-12, phytosterols 2, the esters of sterols to 1.7. The influence of the latter in the form of amaranth oil, which is contained in a medicinal product in a dose of 62.5-250 mg/kg of body weight were studied in the development of experimental toxic hepatitis (ATG).

Drug comparison drug action was a means of plant origin, namely, Essentiale dose of 80 mg/kg

The experiment was performed on 30 white rats-males weighing 180-250 g, contained in standard vivarium conditions Voronezh state medical Academy named. N.N. Burdenko.

The experiments were conducted in the spring. In each series of experiments used rats of the same age group, pre-starving 18-20 hours a Seed crystal animals was carried out by the introduction of the CCl4for 1 day, which allowed to achieve destruction of the liver parenchyma. Day toxic liver lesion formed by subcutaneous doing 50% solution CCL4on peach oil at a dose of 0.4 ml per 100 g weight of the animal once. In the process of modeling the experimental hepatitis mortality of animals was not observed.

This scheme and the dose of CCl4are sufficient to create a complete biochemical picture of toxic damage of the liver parenchyma, and the possibility to follow the direction of the induced patol the policy process.

Before therapy was assessed by biochemical indexes of blood, as well as histological and histochemical sections of liver, kidneys and adrenal glands, for signs of hepatitis, were determined biochemical parameters: the concentration of malondialdehyde (MDA), AST, Alt, hexose associated with protein and thiol samples.

In intact animals and ATG used germ oil and shells of the seeds of amaranth (hereinafter Amaranth oil)containing phospholipids, Tocopherols, unsaturated fatty acids, squalene, phytosterols and esters of sterols in the following ratio, wt.%: phospholipids 7-9, tocopherol 0,3, unsaturated fatty acids 51,25, squalene 3-12, phytosterols 2, the esters of sterols to 1.7.

The control animals oil was introduced through the mouth with a metal probe in the amount of 62.5, 125 and 250 mg/kg, and the animals of the experimental group for 1 h before the introduction of the CCl4. Slaughter animals in the control and experimental groups was carried out by the method of single-step decapitate under chloroform anesthesia after 1 day from the beginning of the seed, respectively. The study was performed in comparison with a solution Essentiale introduced into the stomach in a dose of 80 mg/kg

Development ATG was confirmed by biochemical and morphological methods. Biochemical study were subjected to serum obtained by centrifugation of whole blood at the tip is of 15 min at 3000 rpm, spectrophotometric method.

For morphological studies, tissue samples of the liver lobe size 3×3×2 mm, as well as kidney, heart, adrenal glands and were fixed in 10% solution of neutral formalin. Paraffin sections 4-5 microns thick were stained with hematoxylin and eosin and van gieson.

After a single injection of CCl4in experimental animals have evolved toxic liver damage, which was confirmed:

- increased activity relative to the original level of enzyme markers of liver cytolysis Alt and AST, respectively 143,1% and 83.5% (p<0,05);

- increase in the serum of abiego bilirubin 63.8%. It says or destruction of red blood cells and hemoglobin or violation in the system of direct transfer of bilirubin from the liver cells into the bile;

the increase in the total hexoses on 19,46%, which indicates inflammation;

- reduction Svobodnye SH-groups of 9.4%, clearly associated with a decrease in activity of peroxidase;

- an increase in the concentration Milanofiori of aldehide compared to the original vicinay 25%, which indicates the activation of GENDER and degradation of polyunsaturated fats reactive forms of oxygen.

A single subcutaneous administration to rats Essentiale dose of 0.085 mg/kg caused in relation to the control:

- reduction in the activity of AST, Alt, almost to ishodnoj Turova, that is, respectively 64,5% and 52.2%;

- increase in serum total bilirubin by 13% compared to the control;

- the reduction of total hexoses by 22.5%.

Morphologically preserved lobular structure of the liver, prevailed fat globular canterburyana form of malnutrition, atomized were observed during the transition to relatively preserved periportal zone.

Table 1 shows the biochemical parameters in hepatitis caused by CCl4on a background of application of different doses of oil allocated by cold pressing of the embryo and membranes seeds of amaranth, in the following ratio, wt.%: phospholipids 7-9, tocopherol 0,3, unsaturated fatty acids 51,25, squalene 3-12, phytosterols 2, the esters of sterols to 1.7.

In the experiments used the red blood cells white outbred rats, which were taken from the tail vein by resection of the tip of the tail. Blood is collected by gravity in a test tube containing 0.1 ml of heparin (5000 IU per 1 ml) based on 1-1,5 ml of blood. The blood was centrifuged for 10 minutes at 3000 rpm Then cleaned of nagoshi internal liquid and washed three times under the same conditions. The pure suspension of erythrocytes was prepared with the use of device PE VI, the concentration was monitored spectrophotometrically. Acid resistance of erythrocytes was evaluated with use what Itanium method of registration acid erythrogram. Hemolysis was induced with 0.1 M hydrochloric acid. Erythrogramma registered after the preliminary oral administration of the substances under study (day one) using a metal probe, designed MD, Professor V.A. Nikolaevskiy. An hour before blood sampling in a/b was introduced CCl4at a dose of 0.4 ml/kg of 50% solution in olive oil. Erythrogramma were assessed by the Kmax, tlat, GSF, G50the data presented in table 2.

Table 2
IndicatorsKmax, otedG, %tlatWithG50, %
control5.777±0.93-2.6±1.3125±45.3256,7±37,9
Amaranth oil, 62.5 mg/kg4.455±0.23; t=2.4; P=0.07-1.25±0.95; t=-1.5; p=0.265±31.2 t=1,87; P=0.13235±8,7; t=0.96; P=0.39
Amaranth oil, 125 mg/kg5.26±0.97; t=0.65; P=0.55-1.2±1.22; t=-1.4, P=0.23 105±30; t=0.6; P=0.56246.7±15.3: t=0.42; P=0.7
Amaranth oil, 250 mg/kg4.4±0.71; t=2.04; P=0.111-2.65±0.3; t=0.01; P=0.99105±21.2; t=0.55; P=0.61230±40; t=0.8; P=0.45
Essentiale N, 80 mg/kg5.89±0.37; t=-0.19; P=0.86-0.48±1.1 t=-2.2; P=0.0980±48.2; t=1.17; P=0.31228.3±2.9; t=1.29, P=0.27
CCl48,114±0.0005; t=-4.4; P=0.01-0.45±0.096, t=-2.9; P=0.0450±17.3; t=2.65; P=0.05205±8.7; t=3.7; P=0.02

The analysis of the Kmaxthat reflects the proportion of red blood cells, simultaneously entering into a stage of actual hemolysis, showed that the rate of hemolysis of erythrocytes main srednestan population of erythrocytes with the introduction of Amaranth oil in doses of 62.5-125-250 mg/kg and Essentiale dose of 80 mg/kg were not significantly different from the control experiment.

However, the preliminary introduction of Amaranth oil in doses of 62.5-600-1200 mg/kg on average contributes to the reduction of Kmaxrelative to the control experiment 22.9%, 8.8%and 23.8 per cent, respectively, in the case of the use Essentiale before the introduction of the CCl4share erytrocyte is, at the same time entering into a stage of actual hemolysis, were higher by 2% compared with erythrocytes of animals not treated with the investigated substances. A single on/b introduction CCl4at a dose of 0.4 ml/kg 1 hour before check erythrogram was accompanied by a significant increase in the rate of hemolysis of erythrocytes by srednestan population by 40.6% relative to the control experiment.

Thus, on the basis of in vivo experiments found that CCl4at the specified method and dose of injection causes damage to the cytoplasmic membrane the main population of cells, possibly by increasing oxidative processes in the cell membrane. The preliminary introduction of Essentiale and Amaranth oil has helped to reduce the damaging effects of CCl4possibly due to the proven antioxidant properties of these pharmacologically active substances, which are implemented by increasing the threshold permeability of membranes for hydrogen ions on the application of CCl4separately.

With the introduction of amaranth oil in doses of 62.5-125-250 mg/kg the value of GSFwas not significantly different from the control. However, at doses of 62.5-125 mg/kg, the proportion of spherocytes average was two times less than control at doses of 250 mg/kg practically equal.

With the introduction of Essentiale in a dose of 80 mg/kg tended to decrease GSFbol is e expressed (P=0,09), as evidenced by the decrease in the proportion of red blood cells, while in the stage of spherocytosis 84.6% relative to control. With the introduction of the CCl4GSFsignificantly (P<0,05) less than control at 73%.

Thus, CCl4reduces the acid resistance of erythrocytes by niskalaukaus population by 73% relative to the control, and preliminary application of amaranth oil substantially prevents a direct damaging effect CCl4on the membrane of the "old" erythrocytes.

tlatacid hemolysis, reflecting the duration of latent period of hemolysis, when using Amaranth oil at doses of 62.5-125-250 mg/kg and Essentiale dose of 80 mg/kg was not significantly different from control and was about on par with control. However, in a/b introduction CCl4at a dose of 0.4 ml/kg, was accompanied by a reduction of the latency time of hemolysis by 60% relative to control.

Thus, the use of CCl4separately causes an acceleration of the transition of the erythrocyte stage of ferulacea the stage of actual hemolysis is more than two times, while the preliminary introduction of amaranth oil and Essentiale slow down the development stage actually hemolysis and bring it back to the control levels.

Analysis of the time of occurrence of 50% hemolysis (G50,%) showed that amaranth wt is about all doses and Essentiale cause slowing of the development of hemolysis of erythrocytes main srednestan population regarding the application of CCl 4separately. This is reflected in the fact that pre intragastric administration of Amaranth oil in doses of 62.5-125-250 mg/kg and Essentiale dose of 80 mg/kg before applying CCl4the value of the studied parameter and almost did not differ significantly from those of the control group of animals, which was administered intragastrically once a physiological NaCl solution, and the introduction of CCl4caused the reduction of the time 50% of hemolysis 20.1% relative to control.

Based on the above it is established that the conditions of a particular experiment in vivo prophylactic use of amaranth oil in doses of 62.5-125-250 mg/kg and Essentiale dose of 80 mg/kg increases the acid resistance of erythrocytes in the background of the introduction of the CCl4by weakening the direct damaging effect of the latter on erythrocyte membrane. This is due to the proven antioxidant properties of these substances. In addition, it has to be interaction with the protein-lipid complexes of cell membranes, which leads to changes in the structural and functional properties of erythrocyte membranes and implemented to reduce the permeability of membranes to ions N+regarding the application of CCl4separately.

A therapeutic agent for the prevention and treatment of chronic liver diseases, presented is a shining amaranth oil, characterized in that the latter is obtained by cold pressing of the embryo and membranes seeds of amaranth, which is used at a dose of 62.5-250 mg/kg of body weight.



 

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