RU2538666C2 - Preparation for normalisation of lipid peroxidation processes in animals - Google Patents

Abstract

FIELD: veterinary medicine.

SUBSTANCE: preparation for normalisation of lipid peroxidation processes in animals comprises, wt %: 2-ethyl-6-methyl-3-hydroxypyridine succinate 20.0-30.0, ascorbic acid 5.0-7.0, selenium (Se°) 0.3-0.5, polyvinylpyrrolidone 3.0-5.0, water for injection - the rest.

EFFECT: antioxidant effect, prevention of excessive formation of lipid peroxidation products, high efficiency in normalisation of functioning of the system of antioxidant protection of animals and the process of lipid peroxidation, low toxicity and ease of administration.

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Description

FIELD OF THE INVENTION

The invention relates to veterinary pharmacy, in particular to preparations for the normalization of lipid peroxidation processes in the body of animals with antioxidant properties.

State of the art

Known drug for the correction of stress conditions in animals, providing increased antioxidant and anti-stress effect, reducing the cost of its manufacture, which contains selenium in the nanoscale state and zero valency (Se °), ascorbic acid and water for injection in the following ratio of components, wt.% :

selenium (Se °) 0.001-1.0 ascorbic acid 5.0-20.0 water for injections rest

(See Pat. RU No. 2409347, class A61K 9/08, A61K 33/04, A61P 25/00, published on January 20, 2011)

The disadvantage of this drug is the lack of a specific synthetic antioxidant component in its composition and, due to this, an insufficient antioxidant effect and a low effect of normalizing lipid peroxidation processes.

Known pharmaceutical composition with 2-ethyl-6-methyl-3-hydroxypyridine succinate for oral administration and a method for its preparation, containing mainly 2-ethyl-6-methyl-3-hydroxypyridine succinate as an active component and excipients: lactose monohydrate, sodium carboxymethyl cellulose and / or collidone 30, magnesium stearate, Opadry II white, which can be used in medicine or veterinary medicine as an oral medication for 2-ethyl-6-methyl-3-hydroxypyridine succinate for prevention azlichnyh diseases and pathological conditions of humans or animals. (See Pat. RU No. 2444359, class A61K 31/4425, A61K 9/20, publ. 03/10/2012)

The disadvantage of this drug is its dosage form in the form of tablets, which cannot provide the onset of the maximum effect and is inconvenient and time-consuming for animals, does not provide an accurate dosage and achieve the necessary therapeutic concentration in the body, as well as its increased cost.

The closest in technical essence and the achieved positive effect and adopted by the authors for the prototype is an antioxidant and antihypoxic drug based on 2-ethyl-6-methyl-3-hydroxypyridine succinate in the form of an infusion solution, including 0.5-10.0 6-methyl- 2-ethyl-3-hydroxypyridine succinate, sodium chloride, sodium hydroxide or hydrochloric acid and water for injection with a certain ratio of components, g / l, intended for the treatment of acute and chronic cerebrovascular disorders ischemic and hemorrhagic of a second nature, acute myocardial infarction, prevention of “reperfusion syndrome”, unstable angina pectoris, acute and chronic pancreatitis.

(See Pat. RU No. 2398583, CL A61K 31/4412, A61K 33/14, A61P 39/06, A61K 47/02, published on 09/10/2010)

The disadvantage of this drug is increased toxicity and insufficient effect of normalization of lipid peroxidation processes in animals.

Disclosure of invention

The objective of the invention is to develop a drug for the normalization of lipid peroxidation processes in animals, which has a pronounced antioxidant effect, low toxicity, high efficiency in normalizing lipid peroxidation processes, ease of administration and dosage.

The technical result that can be achieved using the present invention is reduced to antioxidant action, prevention of excessive formation of lipid peroxidation products, high efficiency in normalizing the functioning of the animal antioxidant defense system and lipid peroxidation process, low toxicity and ease of administration and dosage.

The technical result is achieved using a preparation for normalizing lipid peroxidation processes in animals, including 2-ethyl-6-methyl-3-hydroxypyridine succinate, ascorbic acid, selenium in zero valency and nanoscale state (Se °), polyvinylpyrrolidone and water for injection when the following ratio of components, wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 20.0-30.0 ascorbic acid 5.0-7.0 selenium (Se °) 0.3-0.5 polyvinylpyrrolidone 3.0-5.0 water for injections rest

The essence of the preparation for the normalization of lipid peroxidation in animals is as follows: starting materials in wt.%, Namely 2-ethyl-6-methyl-3-hydroxypyridine succinate 20.0-30.0, ascorbic acid 5.0- 7.0, selenium (Se °) 0.3-0.5, polyvinylpyrrolidone 3.0-5.0, water for injection, the rest is mixed under aseptic conditions and packaged.

With an imbalance between the intensity of free radical reactions and antioxidant defense, free radical pathology is observed, which is an integral part of the pathogenesis of many diseases of farm animals and poultry (See Karmoliev R.Kh. // Agricultural Biology. - 2002. - No. 2. - C 19-28) .

A defect in the course of the process of free radical lipid peroxidation can significantly reduce the body's resistance to the effects of adverse environmental and internal factors on it, create the prerequisites for the formation, accelerated development and aggravation of the severity of various diseases of vital organs: lungs, heart, liver, kidneys, etc. (See Shatilov A.V., Bogdanova O.G., Korobov A.V. // Veterinary pathology. - 2006. - No. 2 (21). - S.207-211).

The system "lipid peroxidation - antioxidant defense of the body" is actually responsible for the functioning of all organs and systems and maintaining the living standards of the whole organism. If one or more links of this chain is violated, various diseases of the heart, liver, lungs, kidneys, and other organs and systems of the body occur in the system. What is expressed by the corresponding clinical signs, changes in the biochemical composition of blood and indicators of the antioxidant activity of the body (See Shatilov A.V., Bogdanova O.G., Korobov A.V. // Veterinary pathology. - 2006. - No. 2 (21) - S.207-211).

Enzymatic (superoxide dismutase, catalase, peroxidase, glutathione peroxidase - transferase, ceruloplasmin) and non-enzymatic (tocopherol, retinol, ascorbic acid, selenium) mechanisms of regulation of the body's antioxidant defense system provide control over the level of free radicals and molecular products of lipid peroxidation (see A. E., Borisova M.N. // Veterinary medicine. - 2007. - No. 5. - P.25-28).

Antioxidants are used in medical practice in connection with their ability to inhibit lipid peroxidation, stabilize the structure and improve the functions of cell membranes and thus create conditions for homeostasis during interactions of pathogenic factors on the body (see Burkov V.I., Kolesnichenko I.S., Melnichenko V.I. // Veterinary Medicine. - 2003. - No. 10. - S.52-53).

In medical practice, the drug Mexidol is used, which by its chemical nature is 2-ethyl-6-methyl-3-hydroxypyridine succinate, which is characterized by pronounced antioxidative activity and restricts lipid peroxidation in heterogeneous stress states and pathological processes (See Volchegorsky I. A., M.G. Moskvicheva, E.N. Chashchina // Farmateka. - 2007. - No. 20. - P.76-79). This drug has pronounced antihypoxic properties, promotes aerobic oxidation of glucose in acute stress, and prevents the accumulation of lactate both in the central nervous system and in peripheral organs (See Volchegorsky I.A., M.G. Moskvicheva, E.N. Chashchina / / Pharmateka. - 2007. - No. 20. - P.76-79; Devyatkina T.A., Lutsenko R.V., Vazhnichaya E.M. et al. // Questions of medical chemistry. - 2000. - V. 46 . - No. 2. - S.246-249).

T.A. Devyatkina et al. studied the effect of 3-hydroxy-6-methyl-2-ethylpyridine succinate on the carbohydrate content and lipid peroxidation in the liver of white mice under acute stress and established its protective effect on peroxidation processes and the ability to normalize glycogen content (see Devyatkina T.A., Lutsenko R.V., Vazhnichaya E.M. et al. // Questions of Medical Chemistry. - 1999. - T. 45. - No. 3. - S.246-249).

The prophylactic administration of this compound has a protective effect on liver damage under conditions of acute immobilization stress in rats, namely it inhibits lipid peroxidation and increases the antioxidant defense in the organ, normalizes the activity of hepatocyte damage marker enzymes, also normalizes serum bilirubin content and improves detoxification function liver test with bromosulfalein (See Lutsenko R.V., Vazhnichaya E.M. Devyatkina TA. // Experimental and Clinical Pharmacologist I -. 2003. - №3 -. S.56-58).

When studying the antiradical activity, it was shown that the group of compounds of amino and oxymethyl derivatives of 3-hydroxypyridine has a pronounced antiradical activity, one of the substances with the highest activity is 2-ethyl-6-methyl-3-hydroxypyridine (See. Chechet I.V. , Chechet O.Yu., Kuzin VB // Nizhny Novgorod Medical Journal. - 2006. - No. 7. - S.93-99).

Ascorbic acid takes part in redox processes, regulation of carbohydrate metabolism, blood coagulation, stimulation of hematopoiesis, the formation of steroid hormones of the adrenal glands, normalization of capillary permeability (See Melnichenko VI // Veterinary clinic. - 2006. - No. 7. - C.3-5.).

As a redox system, ascorbic acid catalyzes many tissue metabolism reactions (participation in biological oxidation, reduction of disulfide bonds in proteins to sulfhydryl groups). It has functional connections with other vitamins (A, E, B ₁ , B ₂ , B ₁₂ ) (See Khazipov N.Z., Askarova A.N. // Animal Biochemistry. - Kazan, 1999. - 286 p.).

Ascorbic acid is able to oxidize to dehydroascorbic acid and, thus, together with it it represents a redox system that loses and attaches electrons and protons. This vitamin activates the action of enzymes: arginase, amylase, intracellular proteases, participates in the formation of steroid hormones of the adrenal glands and deoxyribonucleic acid, participates in the regulation of carbohydrate metabolism, blood coagulation, hematopoiesis stimulation, amino acid metabolism (tyrosine, tryptophan). Of great importance is given to ascorbic acid as an important biological antioxidant, which along with vitamin E prevents the accumulation of peroxides and thereby ensures the integrity of cell structures. (See Karpenko L.Yu. // The Role of Vitamins and Minerals in the Metabolism of Small Pets. - SPb, Publishing House SPbGAVM. - 2005. - 67 p.).

Currently, selenium, along with vitamins A, E, and C, is considered one of the four major components of the non-enzymatic pathway of the antioxidant-antiradical system of body protection (See Maistrov V.I., Galochkina V.P., Schevelev N.S. // Agricultural Biology . - 2006. - No. 2. - S.64-68).

Selenium is found in all organs and tissues, has an antioxidant effect, stimulates the growth and development of the body, participates in the interaction of proteins, enzymes, photochemical reactions. To a certain extent, it makes up for the lack of tocopherol, is a part of amino acids, and participates in the synthesis of proteins (See Kleimenov R. // Dairy and beef cattle breeding. - 2004. - No. 6. - P.18-20).

The biochemical functions of selenium are very similar to those of vitamin E. Selenium and vitamin E act together and have antioxidant ability (See Diplock A.T. // American journal of clinical nutrion. - 1991. - Vol.53. - P.189-193 ) One atom of selenium is able to replace 700-1000 molecules of vitamin E. The antioxidant activity of selenium-containing proteins is 500 times higher than that of vitamin E (See Evdokimov P.V. // Vitamins, trace elements, biostimulants and antibiotics in animal husbandry and veterinary medicine. - L. , 1974.- 217 p.).

Brief description of drawings and other materials

Figure 1 is given a drug for the normalization of lipid peroxidation processes in animals. Acute toxicity of the drug to normalize lipid peroxidation in animals, table 1.

In Fig.2, the same graphic representation of acute toxicity for white mice of the drug to normalize the processes of lipid peroxidation in animals, Fig. 1.

Figure 3, the same, a graphical representation of acute toxicity for white rats of the drug to normalize the processes of lipid peroxidation in animals, Fig. 2.

Figure 4, the same, the activity of antioxidant enzymes in the blood of rabbits, table 2.

Figure 5, the same, the activity of antioxidant enzymes and the level of reduced glutathione in the blood of sheep, table 3.

Figure 6, the same, the concentration of peroxidation products in the blood of sheep, table 4.

The implementation of the invention

Examples of specific performance of obtaining and testing the drug

to normalize lipid peroxidation in animals.

Example 1

A preparation for normalizing lipid peroxidation processes in animals is prepared by mixing the components under aseptic conditions and dissolving in water for injection in the following ratio of components, wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 15.0 ascorbic acid 4.0 selenium (Se °) 0.2 polyvinylpyrrolidone 2.0 water for injections rest

The preparation obtained, when administered to rabbits, had a significant effect on the activity of antioxidant system enzymes, which resulted in an increase in catalase activity by 24.2%, superoxide dismutase by 18.6%, and glutathione peroxidase by 21.1%, but there was an insufficient decrease in the concentration of peroxidation products lipids, so the concentration of malondialdehyde decreased by 12.8%, diene conjugates by 16.4%.

Example 2

Carried out analogously to example 1, but take the following ratio of components in wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 20.0, ascorbic acid 5.0, selenium (Se °) 0.3, polyvinylpyrrolidone 3.0, water for injection, the rest. The introduction of the drug to rabbits increased the activity of antioxidant enzymes, in particular catalase by 31.3%, superoxide dismutase by 28.1% and glutathione peroxidase by 34.1%. The concentration of peroxidation products decreased and was within the physiological norm, in contrast to example 1, namely, the level of diene conjugates decreased by 36.9% and malondialdehyde by 32.4%. In addition, positive dynamics was observed with respect to hematological parameters and the level of total protein.

Example 3

Carried out analogously to example 1, but take the following ratio of components, wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 25.0 ascorbic acid 6.0 selenium (Se °) 0.4 polyvinylpyrrolidone 4.0 water for injections rest

When the preparation was administered to rabbits, it was noted that the activity of catalase increased by 33.1%, superoxide dismutase by 31.4% and glutathione peroxidase by 35.5%, and the concentration of lipid peroxidation products decreased and was within the physiological norm, a decrease in the concentration of diene conjugates by 37.4% and malondialdehyde by 33.8%. In general, there is a positive dynamics of changes in biochemical and hematological parameters.

Example 4

Carried out analogously to example 1, but take the following ratio of components, wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 30,0 ascorbic acid 7.0 selenium (Se °) 0.5 polyvinylpyrrolidone 5,0 water for injections rest

The introduction of the drug to rabbits increased the activity of catalase by 35.8%, superoxide dismutase by 32.7% and glutathione peroxidase by 37.7%. The level of lipid peroxidation products in the blood decreased significantly. So, the concentration of diene conjugates in the experimental group was lower by 41.2% and malondialdehyde by 36.5%. A positive effect on the dynamics of the main hematological and biochemical parameters was noted.

Example 5

Carried out analogously to example 1, but take the following ratio of components, wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 35.0 ascorbic acid 8.0 selenium (Se °) 0.6 polyvinylpyrrolidone 6.0 water for injections rest

The resulting preparation, when administered to rabbits, contributed to an increase in the activity of enzymes from the body's antioxidant defense system, catalase by 29.1%, superoxide dismutase by 26.3% and glutathione peroxidase by 30.3%. The concentration of peroxidation products also decreased significantly, but was higher than the upper limits of the physiological norm, while the concentration of malondialdehyde decreased by 12.3%, diene conjugates by 15.1%.

Thus, examples 2, 3 and 4 are the most optimal, since as a result of using the drug, rabbits receive the most significant positive effect of increasing the activity of enzymes from the enzymatic link of the body's antioxidant defense system, in particular catalase from 31.3 to 35.8%, superoxide dismutase from 28.1 to 32.7%, glutathione peroxidase from 34.1 to 37.7%, while there is a decrease in the concentration of lipid peroxidation products in animal blood to physiologically normal values. In the implementation of these examples, a positive effect of the administration of the drug on the main biochemical and hematological parameters, namely, the level of hemoglobin, the number of red blood cells and white blood cells, the level of total protein and glucose, was noted.

To assess the drug to normalize the processes of lipid peroxidation in animals for acute toxicity take the following ratio of components, wt.%:

2-ethyl-6-methyl-3-hydroxypyridine succinate 25.0 ascorbic acid 6.0 selenium (Se °) 0.4 polyvinylpyrrolidone 4.0 water for injections rest

Toxicological studies are carried out on white laboratory mice and laboratory rats, which allows to determine lethal doses, classify the drug according to GOST 12.1.007-76 and classify it as hazard class 4 - low toxic substances (Figs. 1, 2, 3).

The study of the effect of the drug to normalize the processes of lipid peroxidation in animals on the functioning of the body's antioxidant defense system is carried out on rabbits. Twenty animals were divided into two equal groups, taking into account the principle of analogues. In the first group, a drug is administered to normalize lipid peroxidation processes in animals once intramuscularly at a dose of 5 mg / kg body weight. Rabbits from the second group of drugs do not receive and serve as control. All animals are in the same conditions of keeping and feeding. Blood for analysis is taken from the ear vein before drug administration, 3 and 7 days after drug administration. In the blood, the activity of catalase, peroxidase and glutathione peroxidase is determined.

As a result of the studies, it was found that the use of the drug to normalize lipid peroxidation processes in animals to rabbits had a positive effect on the dynamics of the activity of antioxidant enzymes (figure 4). So, for seven days of the experiment, the activity of catalase in the blood of animals from the first group increased by 25.5%, and in the control group decreased by 3.2%, peroxidase activity in the first group increased by 14.5%, and in the second - by 3.4%, the activity of glutathione peroxidase in the first group increased by 26.6%, while in the control group only 2.3%.

The effectiveness of the drug for the normalization of lipid peroxidation processes in animals was studied in two-year-old sheep of the North Caucasian meat and wool breed. The essence of the experiment was that in the spring, before switching to grazing, a planned deworming is carried out in sheep. Twenty heads of queens were allocated from the flock and divided into two equal groups, taking into account the principle of analogues. As an anthelmintic, animals are given orally albendazole suspension of 10% (manufacturer Invesa, Spain) at the rate of 1 ml per 10 kg of animal weight. In the first group of sheep, three days after the use of albendazole, the suspension is administered once intramuscularly to normalize the processes of lipid peroxidation in animals at the rate of 5 mg / kg body weight. In the second group, the drug is not administered to normalize lipid peroxidation in animals - it was a control. Blood is taken from animals from the jugular vein before deworming, before administration of the drug to normalize lipid peroxidation processes in animals, five and ten days after administration of the drug to normalize lipid peroxidation processes in animals. In blood stabilized with heparin, the activity of catalase, peroxidase, superoxide dismutase and glutathione peroxidase, the level of reduced glutathione, the concentration of malondialdehyde, diene conjugates and fluorescent Schiff bases are determined.

The deworming caused a decrease in the activity of the enzymatic component of the animal antioxidant defense system (Fig. 5). In particular, the activity of catalase in the blood for three days decreased in the first group by 4.6%, and in the second by 3.6%. Further dynamics in this indicator significantly varied between groups, for example, in the blood of sheep, which were injected with the drug to normalize lipid peroxidation, the catalase activity increased in animals, after five days the increase was 11.0%, and by the end of the experiment 24.5%, while in the blood of control animals for five days there was a decrease of another 2.3%, and by the end of the experiment a slight increase of only 2.9%.

Peroxidase activity in the blood decreased slightly with the use of anthelmintic in the first group by 2.2%, in the second by 1.1%. In total, since the drug was used to normalize lipid peroxidation processes in animals, sheep in the first group had an increase in blood peroxidase activity of 12.4%, and in the second 4.4%.

After deworming, the activity of superoxide dismutase in the blood decreased in the first group by 7.3%, and in the second by 9.2%, which before the end of the experiment then increased in the first group by 27.1%, and in the second by 6.4%.

The activity of glutathione peroxidase in the blood after the use of albendazole decreased in the first group by 4.2%, in the second by 3.9%. By the end of the experiment, the activity of glutathione peroxidase in the blood of control animals decreased by another 5.41%, and in the blood of sheep from the first group increased by 28.4%.

The most significant deworming effect affected the level of reduced glutathione, which decreased in the blood of sheep from the first group by 9.1%, and in the blood of sheep from the second group by 10.8%. Ten days after the drug was used to normalize lipid peroxidation processes in animals of sheep from the first group, the level of reduced glutathione was restored to the value that was at the time the experiment began, and in the control group for this period of time this indicator decreased by another 6.9% .

When analyzing the dynamics of the concentration of lipid peroxidation products in the blood of sheep, it was noted that the introduction of anthelmintic led to an increase in the level of malondialdehyde, diene conjugates and Schiff's fluorescent bases (Fig. 6). So, the concentration of malondialdehyde in the blood within three days after the administration of albendazole increased by 10.3% in the first group, in the second by 8.6%, but after using the drug to normalize lipid peroxidation in animals in the first group, after five days there was a decrease in this indicator by 23.5%, and from the introduction of the proposed drug until the end of the experiment, the decrease was 42.6%, while in the control second group the concentration increased by another 6.4%.

After deworming, the concentration of diene conjugates increased by 12.1% in the first group and by 12.8% in the second, but until the end of the experiment in the first group it decreased by 39.4%, and in the second by only 5.2%.

The level of Schiff's fluorescent bases increased slightly after the introduction of anthelmintic, in both groups within 6%. From the day the drug was administered to normalize lipid peroxidation in animals and until the end of the experiment, the concentration of fluorescent Schiff bases in the first group decreased by 18.5%, and in the second group increased by 3.8%.

The results of the experiment indicate that the use of the drug to normalize lipid peroxidation processes in animals is an effective way to protect the body from the damaging effects of excessively formed toxic lipid peroxidation products. This happens due to the stimulation of the activity of the enzymatic link of the antioxidant defense system of the body and the direct neutralizing effect of the components of the present invention.

The advantages of the proposed drug is that it contains in the complex of 2-ethyl-6-methyl-3-hydroxypyridine succinate with powerful natural antioxidants - ascorbic acid and selenium taken in a state of zero valency and nanoscale state, which significantly increases its effectiveness, and stable an aqueous solution obtained due to the presence of polyvinylpyrrolidone makes it convenient for animal dosage and use.

The invention in comparison with the prototype and other technical solutions has the following technical advantages:

- increase antioxidant action;

- the most pronounced effect of reducing the concentration of by-products of lipid peroxidation;

- low toxicity;

- complex action;

- increase in biological activity;

- ease of administration and dosage.

Claims (1)

A preparation for normalizing lipid peroxidation processes in animals, including 2-ethyl-6-methyl-3-hydroxypyridine succinate and water for injection, characterized in that it additionally contains ascorbic acid, selenium in zero valency and nanoscale state (Se °) and polyvinylpyrrolidone in the following ratio, wt.%:
2-ethyl-6-methyl-3-hydroxypyridine succinate 20.0-30.0 ascorbic acid 5.0-7.0 selenium (Se °) 0.3-0.5 polyvinylpyrrolidone 3.0-5.0 water for injections rest

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