RU2750798C1 - Antinematodosis composition for veterinary medicine and method for its preparation - Google Patents

Abstract

FIELD: veterinary medicine.

SUBSTANCE: invention relates to an antinematodosis composition and a method for its preparation. The antinematodosis composition contains 5.7-dinitro-4.6-bis(3-nitrophenylamino)benzofuroxane, n-hexadecyltriphenylphosphonium bromide, glucose, 4.6-bis (4-hydroxyphenylamino) – 5.7-dinitrobenzofuroxane, activated carbon, polyvinylpyrrolidone and polyethylene glycol 1500, with the following ratio of components, weight percentage: 5.7-dinitro-4.6-bis(3-nitrophenylamino)benzofuroxane-5; 4.6-bis (4-hydroxyphenylamino) – 5.7-dinitrobenzofuroxane 5; n-hexadecyltriphenylphosphonium bromide 1; activated carbon 10; polyvinylpyrrolidone 10; polyethylene glycol 1500 10; glucose 9. The method for obtaining an antinematodosis composition consists in that the first mixture is prepared, for which 9, parts by weight, glucose is ground, n-hexadecyltriphenylphosphonium bromide, 1, is added with stirring; then the second mixture is prepared, for which 10, in weight parts, polyethylene glycol 1500 is melted at 65 °C, 5, weight parts, 5.7-dinitro-4.6-bis(3-nitrophenylamino)benzofuroxane, 5, weight parts, 4.6-bis (4-hydroxyphenylamino) – 5.7-dinitrobenzofuroxane; then both mixtures are mixed at 65 °C, 10 weight parts of activated carbon and 10 weight parts of polyvinylpyrrolidone are added, the mixture is mixed until a homogeneous mass is obtained, passed through a sieve with a pore diameter of 2 mm, the resulting granules are dried at room temperature and packaged.

EFFECT: proposed antinematodosis composition can be used as an antinematodosis drug in veterinary medicine.

2 cl, 4 dwg

Description

The invention relates to veterinary medicine, in particular, concerns the development of a composition intended for the treatment of agricultural and domestic animals against helminthiasis caused by worms of the nematode series, and a method for its preparation.

Nematodes - roundworms (roundworms, esophagostomas, hookworms, toxocaras, whipworms, pinworms, etc.) are the most common group of helminths and the cause of the main parasitic diseases of agricultural and domestic animals, including birds, which cause significant damage to the economy and pose a danger to humans ... This is due to the high degree of susceptibility to the disease, to which, even after cure, immunity is not developed. The development of drugs for the prevention and treatment of nematodoses affecting various links of the pathological process is an urgent task in pharmacy.

Known anthelmintic synthetic drugs in terms of chemical structure are divided into derivatives of ethanolamine, benzimidazole, salicylamide, piperazine salts [Mashkovsky M.D. Medicines. - M .: New Wave. T 2. - 2004; D.K. Chervyakov and other Medicines in veterinary medicine. - M. Kolos, 1977; Pat. RF 2242976, publ. 27.12.2004; Patent RF 2211835, publ. 10.09.2003]. Also known carbonyloxycyanomethyl derivatives [US Pat. RF 2296747, publ. 27.10.2005], milbemycin derivatives [US Pat. RF 2226531, publ. 10.04.2004; RF application 2004115744, publ. 10.04.2005], derivatives of tetrahydrofuroisoxazole [Patent RF 1586154, publ. 20.12.2006] as antiparasitic agents. Known composite anthelmintic agents, including fenbendazole, sulfadimezin, sulfur and zeolite [Patent RF 2302864, publ.20.07.2007], closantel, novocaine, polyvinylpyrrolidone [US Pat. RF 2278661, publ. 16.02.2004], albendazole, copper sulfate and cellulose acetate [US Pat. RF 2195280, publ. 27.12.2002].

The disadvantages of known drugs are:

- side high toxic effect on the liver, nervous system, decreased immunity;

- allergic phenomena;

- low efficiency of the intended use;

- high therapeutic doses;

- long time of use.

Derivatives of benzofuroxans are known from the prior art as biologically active compounds with a wide spectrum of biological activity, for example, they are known to be used for the treatment of angina pectoris [RF Application 2000132204, publ. 11/10/2002; RF application 2000132206, publ. 20.10.2002]. Also identified them (derivatives of benzofuroxans) acaricidal and bactericidal [US Pat. RF No. 2255935], fungicidal [US Pat. RF No. 2032678], and anti-nematode activity [US Pat. RF No. 2404769, US Pat. RF No. 2404975].

The introduction of components with antimicrobial, anti-inflammatory, prolonged, antiallergenic, detoxifying effect into the composition of anthelmintic (antinematode) drugs, according to the applicant, will lead to the creation of a new drug that expands the arsenal of known agents for this purpose.

In addition, the composition should have an anti-inflammatory and sedative effect on the intestinal mucosa in order to avoid exacerbations and, at the same time, prolonged action. In this case, the composition should act as a soft sorbent for binding toxins released by dying parasites.

The role of the antimicrobial and anti-inflammatory component in the composition of the claimed composition is assigned to 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan, which suppresses microbial infection accompanying inflammatory processes in helminthic invasions.

The role of the prolongator, as a component in the composition of the claimed composition, is assigned to activated carbon. The use of activated carbon as an excipient for prolonged release of a drug in the gastrointestinal tract is known (eg, Roivas, L. et al., Methods Find Exp Clin Pharmacol, 16 (2): 125-32, 1994). The prolonged action of activated carbon can be explained by the known fact that adsorption is not a unidirectional process, this process is always accompanied by desorption of the drug from the activated carbon.

Thus, it is known from the studied prior art that activated carbon is a well-established and widespread agent for preventing the absorption of medicinal compounds from the gastrointestinal tract in case of intoxication due to their adsorption and finds medical use as a detoxifying agent due to the high absorbing capacity of poisons and toxins [Dawson, Andrew. // Australian Prescriber. - 1997. - T. 20. - S. 14-16. - doi: 10.18773 /austprescr.1997.008, Water treatment: Handbook. // Ed. S.E.Belikova. M .: Aqua-Term, 2007.— 240]. The absorption capacity of activated carbon is associated with a porous structure with an inner surface of about 500-1500 m ² / g and a surface area, respectively, of BET up to about 5000 m ² / g.

Polyvinylpyrrolidone plays the role of a component with a detoxifying effect in the claimed anti-nematode composition. The mechanism of the therapeutic action of polyvinylpyrrolidone lies in the complexing ability to actively bind toxins formed during the death and destruction of cuticles of a large number of nematodes in the host's body, and to remove them from the body through the intestines. It is important to note that polyvinylpyrrolidone does not enter the bloodstream. Clinical practice has proven that the drug does not damage the mucous membrane of the stomach and intestines (already inflamed from a large number of nematodes), does not accumulate in the body, has practically no contraindications and side effects, that is, it is a completely safe drug. Indications for the use of polyvinylpyrrolidone: toxic forms of acute infectious diseases of the gastrointestinal tract (dysentery, salmonellosis, etc.), food poisoning (food poisoning), intoxication (poisoning) of other origin, exacerbation of chronic enterocolitis (inflammation of the small and large intestine (intestinal) and enteritis inflammation of the small intestine), liver failure [Medicines, ppm. Mashkovsky, Enterodez].

It is known that polyethylene glycol 1500 (hereinafter referred to as PEG 1500) is a polymer of ethylene glycol, white in color and oily or flaky state. PEG 1500 promotes better mixing and dissolution of drug components, enhances the effect of antiseptics, local anesthetics, hydrocortisone, sulfonamides and many others. It is not very susceptible to bacteria and has an anti-inflammatory effect, helps to speed up the healing of wounds (including infected ones), burns, and eliminates irritation. It has no toxic effect on the body and is easily washed off with water [http: // apifitopharm / ua / kremovaja-osnova-peg 1500; British national formulary: BNF 76 (76 ed.). Pharmaceutical Press. 2018. pp. 57-58. ISBN 9780857113382].

The closest in terms of the number of common features and the claimed technical result, selected by the applicant as a prototype in relation to the composition and in relation to the method of its production, is the invention under the patent of the Russian Federation No. 2413513 "Anti-nematode composition based on quaternary phosphonium salt and substituted dinitrobenzofuroxan".

The essence of the prototype in relation to the composition is an antinematode composition containing n-hexadecyltriphenylphosphonium bromide and 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan as active components at a weight ratio of 1:10 and glucose as an excipient ...

The essence of the prototype in relation to the method is the method, which consists in the fact that first the first component of the known composition - n-hexadecyltriphenyl phosphonium bromide is obtained by interaction of a solution of hexadecyl bromide and triphenylphosphine, the reaction mixture is heated for 15 hours at a temperature of 120 C. The precipitate is filtered off and repeatedly washed with ethyl ether from the starting compounds. Then get the second component - 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan by reacting 4,6-dinitro-5,7-dichlorobenzofuroxan in ethyl alcohol and 3-nitroaniline in ethyl alcohol. The reaction mixture is cooled, a bright red crystal mass is separated and recrystallized from an ethanol-acetonitrile mixture. Then, an anti-nematode composition and a drug based on it are prepared as follows: mix and grind glucose, hexadecyltriphenylphosphonium bromide and 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan until a homogeneous red powder is obtained, which is filled with wafers ( closable metered starch containers). An oral dosage form weighing 0.5 g contains 0.025 g of a phosphonium salt, 0.250 g of 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan and 0.225 g of glucose.

The disadvantages of the prototype in comparison with the claimed technical solution in relation to the composition are:

- the absence of antimicrobial and anti-inflammatory action as a result of the absence in the prototype of the antimicrobial and anti-inflammatory component of 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan;

- lack of prolonged action as a result of the absence of activated carbon in the prototype;

- lack of antiallergenic action as a result of the absence of activated carbon in the prototype;

- lack of detoxification effect as a result of the absence of polyvinylpyrrolidone and activated carbon in the prototype;

- the worst mixing and solubility of the components as a result of the absence of polyethylene glycol 1500 in the prototype;

- an inconvenient dosage form of a known composition - a cachet (a large starch container about 10 mm long), as a result of which the preparation is suitable only for large animals, for example, cows, pigs, horses, and is not suitable for small animals, for example, cats, small breeds dogs, minks, polar foxes, rabbits, chickens, ducks, geese.

The disadvantages of the prototype in comparison with the claimed technical solution in relation to the method of obtaining is the lack of such actions as:

- mixing glucose and n-hexadecyltriphenylphosphonium bromide;

- melting polyethylene glycol 1500 and mixing it with 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan and 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan;

- mixing the two mixtures described above;

- adding activated carbon and polyvinylpyrrolidone;

- passing the mixture through a sieve.

Since the method of obtaining an anti-nematode composition according to the prototype is not included in the formula of the prototype, the second independent claim is drawn up without a limiting part.

Thus, the object of the claimed invention is to create a new composition that additionally has antimicrobial, anti-inflammatory, prolonged, antiallergenic, detoxifying effect and meets the requirements for anti-nematode drugs - low toxicity (hazard class 4), no side effects, low therapeutic doses, providing high efficiency of application, short terms of application, which expands the arsenal of known means of the specified purpose.

The objective of the claimed invention is solved by creating a new composition, which is a combination of biologically active substituted nitro derivatives of benzofuroxan - 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan and 4,6-bis (4-hydroxyphenylamino) -5, 7-dinitrobenzofuroxan, n-hexadecyltriphenylphosphonium bromide, activated carbon, polyvinylpyrrolidone, polyethylene glycol 1500 and glucose in a weight ratio of components 5: 5: 1: 10: 10: 10: 9.

The technical result of the claimed technical solution is:

• creation of an anti-nematode composition for veterinary medicine with:

- antimicrobial and anti-inflammatory action by introducing 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan into the composition;

- prolonged release of the composition, which leads to a more effective, but softer and less traumatic cure, by introducing activated carbon into the composition;

- antiallergenic action by introducing activated carbon into the composition;

- detoxification action by introducing polyvinylpyrrolidone and activated carbon into the composition;

- a more convenient dosage form in the form of small granules as a result of passing through a sieve, which expands the range of animals capable of treatment with the claimed composition;

- the best mixing of the components by introducing polyethylene glycol 1500 into the composition;

- improved solubility of the components of 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan and 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan when dissolved in molten polyethylene glycol 1500.

At the same time, the claimed composition is highly effective against nematodes, as well as low therapeutic doses of 0.044 - 0.047 g / kg and a high LD ₅₀ 3500 mg / kg, which together provides an expansion of the arsenal of known agents for this purpose.

• development of a method for obtaining the claimed composition, which, in the applicant's opinion, has novelty and an inventive step, since the claimed sequence of actions was not identified by the applicant from the prior art and the effect of the combination of features on the claimed technical result was not revealed.

The solution of the tasks and the claimed technical result leads to an increase in the therapeutic efficacy of the claimed composition in comparison with the prototype.

The essence of the claimed technical solution is an anti-nematode composition for veterinary medicine containing 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan, n-hexadecyltriphenylphosphonium bromide, glucose, characterized in that it additionally contains 4,6-bis (4 -hydroxyphenylamino) -5,7-dinitrobenzofuroxan, activated carbon, polyvinylpyrrolidone and polyethylene glycol 1500, with the following ratio of components, wt. h .:

5,7-dinitro-4,6-bis (3-nitrophenylamino) benzofuroxan five 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan five n-hexadecyltriphenylphosphonium bromide one Activated carbon 10 polyvinylpyrrolidone 10 polyethylene glycol 1500 10 glucose nine

A method of obtaining an anti-nematode composition according to claim 1, which comprises preparing the first mixture, for which 9 weight parts of glucose are ground, 1 weight part of n-hexadecyltriphenylphosphonium bromide is added with stirring; then a second mixture is prepared, for which it is melted at 65 ° C 10 parts by weight of polyethylene glycol 1500, 5 parts by weight of 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan, 5 parts by weight of 4,6-bis (4-hydroxyphenylamino) -5 , 7-dinitrobenzofuroxan; then both mixtures are mixed at 65 ° C, 10 parts by weight of activated carbon and 10 parts by weight of polyvinylpyrrolidone are added, the mixture is stirred until a homogeneous mass is obtained, passed through a sieve with a pore diameter of 2 mm, the resulting granules are dried at room temperature and packaged.

The claimed technical solution is illustrated in Fig. 1 - Fig. 4.

Figure 1 presents Table 1, which shows the antimicrobial activity of 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan (1% solution in water) in comparison with the known preparations Chlorhexidine, Penicillin, Griseofulvin.

Figure 2 presents Table 2, which shows the characteristic data of the test gilts before treatment.

Figure 3 presents Table 3, which shows the assessment of the antinematode activity of the investigated compositions according to the results of scatological studies of the feces of pigs.

Figure 4 presents Table 4, which provides an assessment of the antimicrobial, anti-inflammatory, prolonged, antiallergenic, detoxification effect of the investigated compositions.

The set tasks and the claimed technical result is achieved by obtaining a composition consisting of 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan, 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan, n- hexadecyltriphenylphosphonium bromide, polyvinylpyrrolidone (Povidone), activated carbon, polyethylene glycol 1500 (PEG, Macrogol) and glucose at a weight ratio of 5: 5: 1: 10: 10: 10: 9.

Next, the applicant provides a detailed description of the pharmaceutical action of the components of the claimed composition.

5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan - generates NO in vivo and causes spastic convulsions in nematodes, which ultimately paralyzes the muscle tissue of nematodes.

4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan - generates NO in vivo and causes spastic convulsions in nematodes, which ultimately paralyzes the muscle tissue of nematodes. At the same time, it suppresses microbial infection that accompanies inflammatory processes in helminthic invasions. Table 1 in Fig. 1 shows the antimicrobial activity of 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan (1% solution in water) in comparison with the known preparations Chlorhexidine, Penicillin, Griseofulvin. From the data given in Table 1, it can be seen that the antimicrobial activity of 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan significantly exceeds the antimicrobial activity of known drugs.

N-hexadecyltriphenylphosphonium bromide - penetrates through the cuticle membranes, which leads to the death of nematodes and pathogenic microflora due to the availability of the host's digestive juices inside the nematode.

Activated carbon provides both antiallergic and detoxifying effect and prolonged action of the claimed composition. Since the anti-nematode composition acts mainly by direct contact with nematodes in the gastrointestinal tract, namely, mainly in the small intestine, it is very important that the dose of the anti-nematode composition remains in the intestine for as long as possible, for a longer time, due to which prolonged action will be provided.

Prolonged release of the claimed composition is achieved by introducing into the composition activated carbon having a porous structure with an inner surface of about 500-1500 m ² / g and a surface area, respectively, of BET up to about 5000 m ² / g. It is known that activated carbon absorbs medicinal compounds from the gastrointestinal tract in case of intoxication due to their adsorption. However, it is also well known that adsorption is not a unidirectional process, since this process is always accompanied by desorption of the drug from the activated carbon, which is used by the applicant to provide a prolonged release of the claimed composition in the gastrointestinal tract.

Polyvinylpyrrolidone provides detoxification properties of the claimed composition due to its complexation ability. The mechanism of the therapeutic action lies in the ability to actively bind toxins formed during the death and destruction of the cuticles of a large number of nematodes in the host's body, and to remove them from the body through the intestines. It is important to note that polyvinylpyrrolidone does not enter the bloodstream. It is known that the drug does not damage the mucous membrane of the stomach and intestines (already inflamed from a large number of nematodes), does not accumulate in the body, has practically no contraindications and side effects, that is, it is a completely safe drug.

Polyethylene glycol 1500 (PEG 1500) provides better mixing and enhances the solubility of hydrophobic furaxan components - 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan and 4,6-bis (4-hydroxyphenylamino) -5,7 -dinitrobenzofuroxan and thickening of the mixture of the components included in the composition, enhancing the anti-nematode action. In addition, it is known that PEG 1500 is not very susceptible to bacteria and has an anti-inflammatory effect, which helps to accelerate the healing of wounds (including those infected), eliminates irritation caused by the action of nematodes. Moreover, it does not have a toxic effect on the body and is easily washed off with water.

Glucose provides taste masking, and also serves to lure and trick nematodes, since their main food is glucose.

At the same time, the applicant believes that the claimed technical solution is not a summation of known technical solutions both in relation to the composition and in relation to the method of its production, for the following reasons.

With regard to composition.

The composition of the composition selected by the applicant and the ratio of its components is not obvious to a person skilled in the art, since the applicant from the studied prior art did not identify compositions in which the claimed components would be simultaneously used in the claimed ratio and in the declared purpose, namely, in the composition of an anti-nematode composition ... In this case, the ratio of the components was selected by the applicant empirically.

With regard to the method.

The identified sources lack the stated sequence of actions.

Further, the applicant describes the receipt of individual components of the claimed composition:

1. N-hexadecyltriphenylphosphonium bromide of formula I

get according to the method described in [Starks Ch.M. Phase transfer catalysis. Chemistry, Catalysts and Applications / Ed. Ch.M.Starks. M .: Chemistry, 1991.- 157s.]:

2. 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan of formula II is obtained according to the method of [L.M. Yusupov, S.Yu. Garmonov, I.M. Zakharov and others. Means of biological protection of multipurpose appointment based on chlorine derivatives of nitrobenzofuroxan. / Bulletin of Kazan Technological University, 2005, No. 1, p. 103-111.]:

3. 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan of formula III is obtained according to the method [US Pat. RF No. 2255935. 5,7-Disubstituted-4,6-dinitro-benzofuroxan of general formula C6N4O6 (R1) 2, which has acaricidal and bactericidal activity. Garipov T.V., Spatlova L.V., Falyakhov I.F., Yusupova L.M.].

4. Activated carbon, for example, BAU-A GOST 6217-74, is a commercial product.

5. Polyvinylpyrrolidone (Povidone), for example, m. 360000, VWR, food additive E 1201, is a commercial product.

6. Polyethylene glycol 1500 (PEG 1500, Macrogol) pharmaceutical, food additive E 1521, is a commercial product.

7. Crystalline glucose medical, for example, according to GOST 975-88 (manufacturer Dnepropetrovsk starch and treacle plant), is a commercial product.

The claimed technical solution is carried out as follows.

Example 1. Synthesis of n-hexadecyltriphenylphosphonium bromide.

To an excess of 6.1 g (0.02 mol) of a hexadecyl bromide solution, 2.62 g (0.01 mol) of triphenylphosphine are added. The reaction mixture is heated for 15 hours at a temperature of 120 ° C. The precipitate that formed was filtered off and repeatedly washed with ethyl ether to remove the starting compounds.

Yield 87%. M.p. 103.4 ° С, T. decomp. 320.9 ° С, δ_R ³¹ = 25.8 ppm

Elemental analysis C ₃₄ H ₄₈ Br, P. Found,%: C 71.96; H 8.47; P 5.83. Calculated,%: C 71.54; H 8.53; P 5.47.

Example 2. Synthesis of 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan.

To 2.95 g (0.01 mol) of 4,6-dinitro-5,7-dichlorobenzofuroxan in 10 ml of ethyl alcohol, 2.76 g (0.02 mol) of m-nitroaniline in 15 ml of ethyl alcohol are added dropwise. After 30 minutes after cooling the reaction mixture, the crystal mass of bright red color is separated and recrystallized from a mixture of ethanol: acetonitrile (1: 0.8).

The yield is 78%. T pl. 226 ° C.

Elemental analysis C ₁₈ H ₁₀ N ₈ O ₁₀ . Found,%: C 43.71; H 1.97; N 22.34. Calculated,%: C 43.37; H 2.01; N 22.51.

Example 3. Synthesis of 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan.

To 2.95 g (0.01 mol) of 4,6-dinitro-5,7-dichlorobenzofuroxan in 10 ml of ethanol, 2.18 g (0.02 mol) of 4-hydroxyaniline in 15 ml of ethanol are added dropwise. After 30 minutes after cooling the reaction mixture, the crystal mass of a bright burgundy color is separated and recrystallized from a mixture of ethanol: acetonitrile (1: 0.8).

The yield is 85%. T pl. 195-196 ° C (from ethyl alcohol).

Elemental analysis C ₁₈ H ₁₂ N ₆ O ₈

Found,%: C 48.9; H 2.69; N 19. Calculated,%: C 49; H 2.7; N 19.

Example 4. Obtaining an anti-nematode composition.

Stage 1. Get the first mixture, for which in a mortar, for example, porcelain, put 9 weight parts (for example, 2.25 g) of glucose and rub it thoroughly by rubbing into the walls of the mortar in order to smooth out the roughness of the walls in order to avoid further loss of small amounts (1 part by weight) of n-hexadecyltriphenylphosphonium bromide. Then, with stirring, add 1 part by weight (for example, 0.25 g) of n-hexadecyltriphenylphosphonium bromide.

Stage 2. Get a second mixture, for which take 10 parts by weight (for example, 2.50 g) of polyethylene glycol 1500, melted at 65 ° C, for example, in a thermal glass. Next, 5 parts by weight (for example, 1.25 g), 5,7-dinitro-4,6-bis (3-nitrophenylamino) benzofuroxan, and 5 parts by weight (for example, 1.25 g) 4.6 -bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan and stir slowly until dissolved.

Stage 3. Next, both mixtures obtained in Stage 1 and Stage 2 are mixed, for example, in a thermal glass, at 65 ° C, add 10 parts by weight (for example, 2.50 g) of activated carbon and 10 parts by weight (for example, 2.50 g) polyvinylpyrrolidone. The mixture is stirred until a homogeneous mass is obtained, passed through a sieve with a pore diameter, for example, 2 mm. The resulting granules are dried at room temperature. Get the claimed composition.

The granules are packaged in sachets of 0.5 g each, and a prepackaged drug is obtained.

An oral dosage of the claimed composition weighing 0.5 g contains:

0.05 g of 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan,

0.05 g 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan,

0.01 g of n-hexadecyltriphenylphosphonium bromide,

0.1 g of activated carbon,

0.1 g of polyvinylpyrrolidone,

0.1 g polyethylene glycol 1500,

0.09 g glucose

with a weight ratio of 5: 5: 1: 10: 10: 10: 9.

Example 4. Study of antinematozoic activity against mixed pig helminthiasis caused by: Ascaris suum ascaris, Oesophagostomum dentatum esophagostomy and Trichocephalus suis whipworm, as well as prolonged, anti-allergenic, detoxifying, anti-inflammatory action of the claimed composition.

Anti-nematode activity, as well as antimicrobial, anti-inflammatory, prolonged, antiallergenic, detoxifying effect of the claimed composition was studied on pigs at OOO Kolos, Tetyushsky district of the Republic of Tatarstan, as well as in the vivarium of the Department of Parasitology and Radiobiology, Kazan State Academy of Veterinary Medicine. Fecal samples were taken from each spontaneously infested mumps with nematodes, which were examined for the presence of nematode eggs using the Fulleborn method.

Based on the results of helminthic examination, 10 gilts of 3-4 months of age (average weight from 23 to 25 kg) were selected and divided into 5 groups, 2 gilts in each.

In the study of the anti-nematode activity, as well as the antimicrobial, anti-inflammatory, prolonged, antiallergenic, detoxification effect of the claimed composition and its components, the preparations were administered to gilts orally in the form of granules weighing 0.5 g with feed 2 times a day - in the morning and in the evening:

in the 1st group, the claimed composition was used, consisting of all seven components;

in the 2nd group, a composition of six components was used, similar to the claimed one, but not containing 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan;

in the 3rd group, a composition of six components was used, similar to the declared one, but not containing activated carbon;

in the 4th group, a composition of six components was used, similar to the claimed one, but not containing polyvinylpyrrolidone;

in group 5, a composition of six components was used, similar to the claimed one, but not containing polyethylene glycol 1500.

The characteristic data of the test gilts before treatment and the dose of drugs for the course of deworming are presented in Table 2 in Fig. 2. As follows from the data in Table 2, in all groups the blood picture was characterized by anemia, leukocytopenia, erythrocytopenia.

The feces of the pigs were examined 1, 2, 3 and 4 weeks after the use of the investigated compositions.

Evaluation of the anti-nematode activity of the investigated compositions according to the results of scatological studies of feces of pigs are presented in Table 3 in Fig. 3.

Evaluation of the antimicrobial, anti-inflammatory, prolonged, anti-allergenic, detoxification effect of the investigated compositions are presented in Table 4 in Fig. 4.

The results presented in Tables 3 and 4 show the following.

In the 1st group, which took the claimed composition of all 7 components, there was almost complete deworming of the pigs - according to the pathologist's conclusion, after 4 weeks, nematodes were not found in the intestine or were found in an insignificant amount (27 pieces per 1 g of feces), which indicates a high anginematic activity of the claimed composition.

Faeces of a clarified color of plasticine consistency appeared once at the end of the second week of treatment in the 1st guinea pig (microscopy revealed the remnants of the cuticle of ascaris) and twice in the 2nd guinea on the 11th and 14th days in the form of cuticle residues with esophagostome and ascaris, respectively.

There is no pathogenic bacterial microflora in the nasal smear, which indicates the antimicrobial effect of the claimed composition.

At the same time, the deworming process in the 1st group took place within 5 days with a further 3-week gradual cleansing of the body from the remains of nematodes and dead eggs, which indicates the prolonged action of the claimed composition.

There was practically no allergy and cough, which indicates the anti-allergenic effect of the claimed composition. A small cough that appeared temporarily for 1-2 days indicated the death of a large number of nematode larvae in the lungs, and weak allergic reactions, noticeable only for a specialist, were associated with the simultaneous death of a large number of nematodes.

Blood counts completely returned to normal within a week, anemia, leukopenia, erythrocytopenia disappeared, which indicates the absence of allergic, intoxication, and inflammatory processes.

Compared with the pigs of the 2nd, 3rd, 4th groups and 5th groups, in the 1st group the pigs after deworming were the most active, frolicked and played, diarrhea and any other side effects from taking the claimed composition was not observed. It should be noted the improvement in the general condition of the pigs and good appetite after taking the claimed composition.

Thus, from the above, we can conclude that the use of the claimed composition, in comparison with the prototype, showed:

- high anti-nematode effect, comparable to the prototype - according to the results of treatment, nematodes were not found;

- antimicrobial action - blood counts returned to normal after treatment, pathogenic cocci were not found in the microbiological contents (swab from the nose of pigs);

- anti-inflammatory effect - leukocytes returned to normal within a week;

- prolonged action - the time and, accordingly, the quality of the cure increased compared to the prototype - the deworming process took place within 5 days with 3 weeks of cleansing compared to 1-2 days with 2 weeks of cleansing in the prototype;

- anti-allergenic action - allergy, in comparison with the prototype, was practically absent;

- detoxification effect - blood counts and the condition of the pigs completely returned to normal within a week.

In the 2nd group, a composition of six components was used, similar to the claimed composition, but not containing 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan, which provides the antimicrobial and anti-inflammatory effect of the claimed composition.

Within 4 weeks, there was an almost complete cleansing of the intestines of the pigs from nematodes and their remains - according to the pathologist, no nematodes were found in the intestines.

Faeces of a clarified plasticine consistency appeared once, on the sixth day, in the 3rd pig (microscopy revealed the remnants of the ascaris cuticle) and in the 4th pig, on the seventh day, in the form of cuticle remnants of the esophagostom, whipworm and ascaris, respectively. Diarrhea and any other side effects of the composition were not observed.

Blood counts returned to normal more slowly than in other pigs, and pathogenic cocci disappeared in nasal swabs for 2 weeks longer, which, according to the applicant, indicates a decrease in the antimicrobial effect of the composition.

At the same time, the process of deworming in the 2nd group took place within 5 days with a further 3-week gradual cleansing of the body from the remains of nematodes and dead eggs, which indicates a prolonged action.

Within 4-6 days, there was a cough, within 3 weeks there was an increased content of leukocytes in the blood, which, according to the applicant, indicates a decrease in the anti-inflammatory effect of the composition.

There were practically no allergic reactions.

Blood counts, except for leukocytes, completely normalized after 2 weeks, which indicates the detoxification effect of the composition.

Thus, in comparison with the pigs of the 1st, 3rd, 4th and 5th groups, the antimicrobial and anti-inflammatory effect of the composition decreased in the 2nd group.

From the above, we can conclude that the use of a composition that does not contain 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan showed:

- the anti-nematode effect is the same as in other groups - according to the results of treatment, nematodes were not found, however, the cleansing of the pigs was slower;

- prolonged action - the time and, accordingly, the quality of the cure is comparable to the claimed composition;

- anti-allergenic action - there was practically no allergy;

- detoxification effect - blood counts, except for leukocytes, and the condition of the pigs are comparable to the claimed composition;

- a decrease in antimicrobial action - within two weeks, pathogenic cocci were found in a nasal swab;

- decrease in anti-inflammatory effect - blood leukocytes returned to normal after 3 weeks compared to 1 week for the claimed composition.

In the 3rd group, a composition of six components was used, similar to the claimed composition, but not containing activated carbon, which provides both antiallergic and detoxification effects, and a prolonged action of the claimed composition.

Within 4 weeks, there was an almost complete cleansing of the intestines of pigs from nematodes and their remains - according to the pathologist's conclusion, nematodes were not found in the intestines or were found in insignificant quantities (35 pieces per 1 g of feces).

Faeces of a clarified plasticine consistency appeared once - on the sixth day in the 5th pig (microscopy revealed the remnants of the cuticle of ascaris) and in the 6th pig on the seventh day in the form of the remains of the cuticles by esophagostom, whipworm and ascaris, respectively. Diarrhea and any other side effects of the composition were not observed.

There is no pathogenic bacterial microflora in the nasal smear, which indicates the antimicrobial effect of the claimed composition.

At the same time, the process of deworming in the 3rd group took place within 1-2 days (compared to 5 days in the 1st group), with a further 2-week gradual cleansing of the body from the remains of nematodes and dead eggs (compared to 3- x weekly in the 1st group), which indicates the absence of prolonged action of the composition that does not contain activated carbon.

In the pigs of the 3rd group, a cough was observed for 2 weeks, which indicates intoxication and allergic processes, strong skin allergic reactions were observed within 3 weeks. This indicates the absence of antiallergenic effect of the composition, which does not contain activated carbon.

Blood counts completely returned to normal after 2 weeks, leukocytes returned to normal in 1-2 weeks, compared to 1 week in group 1, which indicates a decrease in detoxification effect.

Thus, in comparison with the pigs of the 1st, 2nd, 4th and 5th groups, in the 3rd group there was no prolonged effect of the action of the composition, allergies and cough were more pronounced.

From the above, we can conclude that the use of a composition that does not contain activated carbon showed:

- anti-nematodic action - according to the results of treatment, nematodes were not detected or were found in single quantities;

- antimicrobial action - microbiological examination of nasal swabs without pathogens;

- anti-inflammatory effect - leukocytes are comparable to the claimed composition;

- lack of prolonged action in comparison with the claimed composition - the time and, accordingly, the quality of the cure has decreased;

- lack of antiallergenic action - there was a strong allergy and cough compared to the claimed composition;

- decrease in detoxification effect - blood counts compared with the claimed composition returned to normal after 2 weeks compared to 1 week for the claimed composition.

In the 4th group, a composition of six components was used, similar to the claimed one, but not containing polyvinylpyrrolidone, which provides detoxification properties of the claimed composition due to its complexation ability.

Within 4 weeks, there was an almost complete cleansing of the intestines of the pigs from nematodes and their remains - according to the pathologist, no nematodes were found in the intestines.

Faeces of a clarified plasticine consistency appeared once at the end of the second week of treatment in the 7th and 8th pigs (microscopy revealed remnants of the cuticle of ascaris and other nematodes). The pigs after deworming were active and no other side effects of the drug were observed.

At the same time, the process of deworming in the 4th group took place within 5 days with a further 3-week gradual cleansing of the body from the remains of nematodes and dead eggs, which indicates a prolonged action.

There is no pathogenic bacterial microflora in the nasal swab, which indicates antimicrobial effect.

Within 5-6 days, a cough was observed, as well as minor allergic reactions, which indicates intoxication as a result of the death of a large number of nematode larvae in the lungs. At the same time, the blood contained an increased number of leukocytes.

Blood counts completely returned to normal in 2-3 weeks, leukocytes returned to normal in 1-2 weeks, compared to 1 week in the claimed composition, anemia, leukopenia, erythrocytopenia disappeared.

Thus, from the above, we can conclude that the use of a composition that does not contain polyvinylpyrrolidone showed:

- anti-nematodic action - according to the results of treatment, nematodes were not detected or were found in single quantities;

- antimicrobial action - microbiological examination of nasal swabs without pathogens;

- anti-inflammatory effect - blood parameters are comparable to the claimed composition;

- prolonged action - the time and, accordingly, the quality of the cure is comparable to the claimed composition;

- anti-allergenic action - there was practically no allergy;

- a decrease in the detoxification effect in comparison with the claimed composition - blood counts returned to normal within 2-3 weeks compared to 1 week in the claimed composition, a cough was observed.

In the 5th group, a composition of six components was used, similar to the claimed one, but not containing polyethylene glycol 1500, which provides better mixing and solubility of the hydrophobic components of the claimed composition, helps to thicken the mixture of substances included in the composition and enhance its anti-inflammatory effect.

Within 4 weeks, there was an almost complete cleansing of the intestines of the pigs from nematodes and their remains - according to the pathologist, no nematodes were found in the intestines.

Faeces of a clarified plasticine consistency appeared once at the end of the second week of treatment in pigs 9 and 10 (microscopy revealed remnants of the cuticle of ascaris and other nematodes).

There is no pathogenic bacterial microflora in the nasal smear, which indicates the antimicrobial effect of the claimed composition.

The process of deworming in the 5th group took place within 5 days with a further 3-week gradual cleansing of the body from the remains of nematodes and dead eggs, which indicates a prolonged action.

There were practically no allergic reactions.

Blood counts returned to normal after 3 weeks, anemia, leukopenia, erythrocytopenia disappeared.

When preparing a composition that does not contain PEG-1500, there was a deterioration in the mixing and solubility of the components of the composition in comparison with compositions in groups 1-4.

Thus, from the above, we can conclude that the use of a composition that does not contain polyethylene glycol 1500 showed:

- anti-nematodic action - according to the results of treatment, nematodes were not found;

- antimicrobial action - microbiological examination of nasal swabs without pathogens.

- anti-inflammatory effect - leukocytes are comparable to the claimed composition;

- prolonged action - the time and, accordingly, the quality of the cure is comparable to the claimed composition;

- anti-allergenic action - there was practically no allergy;

- detoxification effect - blood counts, except for leukocytes, and the condition of the pigs are comparable to the claimed composition;

- deterioration of mixing and solubility of the components of the composition during its preparation.

Thus, as a result of the study, it was found that the claimed composition has a high anti-nematodic effect at low therapeutic doses of 0.044 - 0.047 g / kg, while piperazine preparations are used at doses of 0.33 g / kg (8 times higher than the dose of the claimed composition) ... At the same time, there are no stool disorders and other signs of intestinal infection.

The level of acute toxicity of the claimed anti-nematode composition LD ₅₀ 3500 mg / kg in mice under conditions of intraperitoneal and oral administration corresponds to the category of "relatively harmless" - IV class of toxicity according to Izmerov [Izmerov N.F., Sanotskiy I.V., Sidorov K.K. ... Parameters of toxicometry of industrial poisons with a single injection (Reference). - M .: Medicine, 1977. - From 196-197].

Moreover, each component of the claimed composition contributes to its action:

- 5,7-dinitro-4,6-bis- (3-nitrophenylamino) benzofuroxan - a component that generates NO in vivo and causes spastic convulsions in nematodes, and as a result paralyzes the muscle tissue of nematodes;

- 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan - a component that generates NO in vivo and causes spastic convulsions of nematodes, and as a result paralyzes the muscle tissue of nematodes, simultaneously exerting antimicrobial and anti-inflammatory effects;

- n-hexadecyltriphenylphosphonium bromide - a component penetrating through the cuticle membranes, leading to the death of nematodes and pathogenic microflora due to the availability of the host's digestive juices inside the nematode (as evidenced by the destruction of the cuticle of nematodes, the plastic-like state of feces and the absence of nematodes in their feces in animals, or the presence of a single amount of nematodes in their feces) ;

- activated carbon provides prolonged action of the claimed composition and its anti-allergenic and detoxification properties;

- polyvinylpyrrolidone - provides detoxification properties of the claimed composition;

- polyethylene glycol 1500 - provides better mixing and solubility of components, thickening of the mixture of substances included in the composition;

- glucose - provides masking of taste and is a necessary "bait" for the helminth in the intestine of the host, as it is the main nutrient of the helminths.

Thus, from the above, we can conclude that the applicant has achieved the assigned tasks and the claimed technical result, which exceeds the technical result of the prototype, namely:

• an anti-nematode composition for veterinary medicine has been created, which includes seven components - 5,7-dinitro-4,6-bis (3-nitrophenylamino) benzofuroxan, 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan, n-hexadecyltriphenylphosphonium bromide , activated carbon, polyvinylpyrrolidone, polyethylene glycol 1500 and glucose at a weight ratio of 5: 5: 1: 10: 10: 10: 9, which has:

- antimicrobial and anti-inflammatory action by introducing 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan;

- prolonged release of the composition, which leads to more effective, but mild and less traumatic healing by introducing activated carbon into the composition;

- antiallergenic action by introducing activated carbon into the composition;

- detoxification action by introducing polyvinylpyrrolidone and activated carbon into the composition;

- improved mixing of components when dissolved in molten polyethylene glycol 1500;

- a more convenient dosage form in the form of small granules as a result of passing through a sieve, which expands the range of animals capable of treatment with the claimed composition.

At the same time, the claimed composition is highly effective against helminths, as well as low therapeutic doses of 0.044 - 0.047 g / kg and a high LD ₅₀ value of 3500 mg / kg, which provides an expansion of the arsenal of known agents for this purpose.

• a method for obtaining the claimed composition has been developed, which has novelty and inventive step, since the claimed composition and sequence of actions have not been identified by the applicant from the prior art and the influence of the combination of features on the claimed technical result has not been revealed.

The solution of the set tasks and the claimed technical result leads to an increase in the therapeutic effectiveness of the claimed composition in comparison with the prototype and expands the arsenal of means for this purpose.

The claimed technical solution meets the "novelty" requirement of patentability for inventions, since at the date of the applicant's submission of application materials from the studied prior art, no sources have been identified that have a set of features identical to the set of features of the claimed technical solution.

The claimed technical solution complies with the "inventive step" patentability requirement for inventions, since the combination of the claimed features provides for obtaining technical results that are not obvious to a specialist, exceeding the technical result of the prototype. The applicant from the investigated prior art has not identified compositions in which the claimed components would be simultaneously used in the claimed ratio and in the declared purpose, namely, as part of an anti-nematode composition. In this case, the ratio of the components was selected by the applicant empirically. The identified sources also lack the sequence of actions of the claimed method, therefore, the claimed method is not obvious to a person skilled in the analyzed field of technology.

The claimed technical solution meets the condition of patentability "industrial applicability" for inventions, since the claimed composition can be obtained by using known components using standard equipment and known techniques.

Claims (3)

1. Antinematode composition for veterinary medicine containing 5,7-dinitro-4,6-bis (3-nitrophenylamino) benzofuroxan, n-hexadecyltriphenylphosphonium bromide, glucose, characterized in that it additionally contains 4,6-bis (4-hydroxyphenylamino) - 5,7-dinitrobenzofuroxan, activated carbon, polyvinylpyrrolidone and polyethylene glycol 1500, with the following ratio of components, wt. h .: 5,7-dinitro-4,6-bis (3-nitrophenylamino) benzofuroxan five 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan five n-hexadecyltriphenylphosphonium bromide one Activated carbon 10 polyvinylpyrrolidone 10 polyethylene glycol 1500 10 glucose nine 2. A method of obtaining an anti-nematode composition according to claim 1, comprising the fact that the first mixture is prepared, for which 9 weight is ground. including glucose, add with stirring 1 weight. including n-hexadecyltriphenylphosphonium bromide; then prepare a second mixture, for which it is melted at 65 ° C 10 wt. including polyethylene glycol 1500, add to it with stirring 5 wt. including 5,7-dinitro-4,6-bis (3-nitrophenylamino) benzofuroxan, 5 wt. including 4,6-bis (4-hydroxyphenylamino) -5,7-dinitrobenzofuroxan; then both mixtures are mixed at 65 ° C, add 10 wt. hours of activated carbon and 10 wt. including polyvinylpyrrolidone, the mixture is stirred until a homogeneous mass is obtained, passed through a sieve with a pore diameter of 2 mm, the resulting granules are dried at room temperature and packaged.

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