Antioxidant, stress- and neuroprotective pharmacological agent potassium comenate

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmaceutics and medicine and concerns using comenic acid potassium salt as a preventive and therapeutic antioxidant, stress- and neuroprotective agent in the amount of 2 to 8 mg per 1 kg of body weight daily on the empty stomach for 3 days.

EFFECT: agent possess high efficacy.

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The invention relates to experimental medicine and pharmacology, namely tools for the prevention and treatment of neurodegenerative diseases caused by oxidative damage to the brain.

Many diseases of the Central nervous system in which there is destructive processes of brain neurons, associated with activation of lipid peroxidation (LPO). Oxidative stress, leading to hyperproductive free radicals (SRS) and the degradation of membranes associated with activation phospholipase hydrolysis plays a major role in the pathogenetic mechanisms of brain ischemia. Cerebral ischemia triggers a cascade of biochemical reactions underlying tissue damage. The basic mechanisms of neuronal damage include the depletion of energy resources in conditions of acidosis, brain tissue, disruption of ion homeostasis, depolarization of cell membranes, accumulation of excitatory amino acids and hyperproduction of reactive oxygen species (ROS). Chronic oxidative stress leads to damage of all classes of biomolecules, including lipids, proteins and nucleic acids. Excess free radicals negatively affecting DNA of neurons in the brain, can cause a change in the genetic code and the death of cells [MS Popova, Induced M. Induction of cell cycle, amyloid-beta and free radicals in fur the change in the development of neurodegenerative processes in the brain // neurochemistry. 2008. V.25. No. 3. S-178; Zozulya Y.A., Baraboi, VA, sutkovoi D.A. Free radical oxidation and antioxidant protection in the pathology of the brain. M: Knowledge. 2000. S.226-221; Smirnova, I.N., Fedorov T.N., Tanashian M.M., Z.A. Suslina Clinical efficacy and antioxidant activity of Mexidol in chronic cerebrovascular diseases // Nervous illness. 2006. No. 1. Pp.33-36; Fedorova so-CALLED. Oxidative stress and protect the brain from ischemic damage: Diss. Dr. Biol. Sciences: 03.00.04: Moscow. 2004. 298 S. RSL OD. 71:05-3/6; Calabrese V., Bates IE, Stella A.M. NO synthase and NO-dependet signal pathways in brain aging and neuredegenerative disorders: the role of oxidant / antioxidant balance // Neurochem. 2000. V.25. P.1315-1341].

Another important factor causing the death of neurons is prolonged stimulation of neurons in the Central nervous system neurotransmitter glutamate. As an excitatory neurotransmitter, glutamate found in most neurons of the brain. Ischemia glutamate is released from the endings of ischemic neurons in the extracellular space. Under normal conditions, neurons and glia cells absorb excess glutamate from the extracellular space. However, ischemic cells are deprived of the necessary energy. In the result of the accumulation of large amounts of glutamate is the saturation of the receptors on neighboring neurons and the damage zone is expanding. Over-stimulation of glutamate Retz is Perov, accompanying hypoxia, ischemia, has a striking effect on the neurons leads to impaired calcium homeostasis. Deferred calcium down-regulation is one of the key signals, resulting in enhanced generation of ROS and the FLOOR after the toxic effect of glutamate, reducing the level of endogenous antioxidants, damage and death of brain cells in ischemic conditions [Suslin., Maksimov M. the Concept of neuroprotection: new opportunities for urgent therapy of ischemic stroke. Symposium of the Institute of neurology RAMS "Treatment of stroke: the state of the problem" // Nervous illness. 2004. No. 3. P.4-7; O. Vergun, Keelan J., Khodorov B.I., M.R. Duchen Glutamate-induced mitochondrial depolarisation and perturbation of calcium homeostasis in cultured rat hippocampal neurons // J. Physiol. 1999. Vol.519. P.451-466; Pinelis VG, Bykova, L.P., Bogachev A.P. Isaev NK, Victorov IV, Khodorov B. I. Toxic effect of glutamate on cultured granular cells of the cerebellum reduces intracellular ATP content. The role of ions of CA2+// Bul. the experimental. Biol. and the honey. 1997. T. No. 2. S.162-164; Nishizawa Y. Glutamate release and neuronal damage in ischemia // Life Sciences. 2001. Vol.69. P.369-381].

Antioxidant defense during oxidative stress, is unable to neutralize the resulting excess ROS. Under these conditions, the apparent feasibility of using antioxidant compounds that can protect brain tissue from damage [Nonaka Sh., Katsube, N. and Chang D.-M. Lithium Protects Rat Cerebellar Granule Cells against Apoptosis Induced by Anticonvulsants, Phenytoin and Carbamazepine // JPET. 1998. V.286. P.539-547].

Now for the prevention and treatment of degenerative disease with ischemic damage applied antioxidants, which in its origin is divided into two main groups: natural and synthetic.

Natural antioxidants: enzymes, proteins, low molecular weight compounds (vitamins E and C, carotenoids, ubiquinone and other).

A group of synthetic antioxidants are synthetic analogues of vitamin E, aromatic phenols and polyphenols (BHT, probucol), emoxipin and Mexidol, some derivatives of barbituric acid and fenotiazina, preparations of iron and zinc, organic acids and their derivatives, some amino acids and derivatives aminosteroids [Korneev A.N., Solovyov EU and other drugs alpha-lipoic acid as a neuroprotective therapy in chronic ischemia of the brain. Directory of outpatient physician. http://old.consilium-medicum.com/media/refer/06_08/76 .shtml]. While the toxicity of some drugs, including barbiturates may cause reversible cognitive disorders - dysmetabolic dementia [Chukhlovina M. Dementia. Diagnostics and treatment. St. Petersburg: 2010. P.60].

Known use as medicines in various connect the deposits of potassium. This is mainly organic potassium salts (potassium orotate, potassium Asparaginate, potassium acetate and inorganic salts of potassium (potassium chloride) [CRU J. Biochemistry, Medical and biological aspects, trans. from French. M., 1979. p.46; Laboratory methods in the clinic, under the editorship of V.V. Menshikov. M., 1987. Pp.261, 263; http://dic.academic.ru/]

Potassium is the major cation of intracellular fluid, is important for various biological processes. It is important for nerve and muscle activity, since it forms an electrolytic environment, which is essential for maintaining osmotic pressure and acid-base balance, for the normal functioning of nerves and muscle cells in the conduction of nerve impulses in nerve and muscle tissue. The main function of potassium is the formation of transmembrane potential and distribution changes of potential on the cell membrane by exchange with sodium ions in a concentration gradient. Potassium is involved in maintaining the electrical activity of the brain, nervous tissue, contraction of skeletal and heart muscle, slows the heart rate and, acting similarly to the wandering nerve, participates in the regulation of cardiac activity, participates in the regulation of blood pressure. It is known that the decrease in the ratio of sodium/potassium by additives in foods potassium reduces the pressure in GI is erenkov. Potassium supplements can reduce the risk of vascular diseases of the brain, and kidney and heart [Rock AV, Rudakov I.A. Bioelements in medicine. M: 2004. 272 C. http://www.adlshop.ru/node/121].

It is known the use of potassium chloride (Kalii chloridum). Potassium is involved in the process of nerve impulses and transmit them to the innervated organs. The introduction of potassium in the body is accompanied by an increase of acetylcholine and excitation of the sympathetic nervous system. Intravenous marked increase in the adrenal glands.

Contraindications: if you are not careful intravenous drug can cause intoxication (paresthesia). In rare cases there may be a paradoxical reaction - increasing the number of extrasystoles. Ingestion may include nausea, vomiting, diarrhea. The use of potassium chloride is contraindicated in disorders of atrioventricular conduction, complete heart block, renal excretory function. In these cases, potassium accumulates in the blood plasma, which can lead to intoxication. There is evidence of severe complications occurring after administration of poorly soluble dosage forms of potassium chloride. In order to avoid possible complications, it is not recommended to use potassium chloride in the form of conventional tablets

It is known the use of the drug asparkam, sod is rasego potassium and other Asparaginate This drug has properties similar to the properties of potassium chloride. However, in contrast, has a much less pronounced local irritant effect on the digestive tract and is better tolerated. This is because tablets "Asparkam" potassium is found in the form of organic salts, which compared with potassium chloride dissociates to a lesser extent [http://www.mordovnik.ru/solkalia. Potassium salts; Small medical encyclopedia. M: Medical encyclopedia. 1991-1996; First aid. M: Great Russian Encyclopedia. 1994; Encyclopedic dictionary of medical terms. M: Soviet encyclopedia. 1982-1984 http://dic.academic.ru].

Closest to the claimed antioxidant neuroprotective tool is kamenova acid (5-hydroxy-γ-pyrone-2-carboxylic).

Kamenova acid has antioxidant and neurotropic properties [Shurygin YA, Kolendo SV, Yugay GA Antioxidant effect of Kamenovo acid. Izvestiya Vuzov. The North Caucasus region. The natural Sciences. 2000. No. 1. P.100-101, Husary LR the Influence of the stress on morpho-functional characteristics of cultured rat neurons. Abstract. Diss. Kida. Biol. Sciences. M 1998; Shurygin YA Drug basis. 2002. 416].

Installed it pronounced growth-stimulating effect on Kul is URS neurons of the cerebral cortex of prenatally stressed animals (Shurygin YA Drug Balis, 2002, 416 S.). Kamenova acid has a mild sedative effect, not causing addiction [RF patent for the invention №2209062, IPC (7) A61K 31/351, A61P 25/20], is a highly effective non-narcotic analgesic agent, without the negative side effects that do not cause dependence and addiction, resulting in long-term pain syndrome [RF patent №2322977 A61K 31/351 (2006.01), A61P 29/02 (2006.01)]. Has antiobscenity, anxiolytic, and antidepressant properties [Panova TI Mechanisms influence Kamenovo acid integrative activity of the brain : Theoretical and experimental medicine. Medicine today and tomorrow. 2005. No. 1. Pp.28-33].

The technical result is the creation of medicines, reducing neuronal death in conditions of oxidative stress when neurogenerative diseases of the brain.

To achieve a technical result, it is proposed to use as pharmacological agents for the prevention and treatment of neurodegenerative diseases caused by oxidative damage to the brain, potassium salt of Kamenovo acid (coment potassium), obtained by mixing solution Kamenovo acid, heated to a temperature of 70-80°C, with a solution of potassium hydroxide in the amounts taken from the calculation of stoichiometric to mn the treatment pH, equal to 4.6. Coment potassium is used in an amount of from 2 to 8 mg per 1 kg of body weight as an antioxidant, stress and neuroprotective agents once daily, on an empty stomach for 3 days.

To obtain the potassium salt Kamenovo acid take 50 ml of 3 mm hot solution of Kamenovo acid (70-80°C) and add, while stirring, 15 ml of a 3 mm solution of potassium hydroxide. The reaction proceeds rapidly, the solution turns yellow. Potassium salt of Kamenovo acid are separated from the solution by distillation of water under vacuum or by evaporation of water from the salt solution in a water bath. The output of comenta potassium is about 90%.

Antioxidant stress and neuroprotective properties of the potassium salt Kamenovo acid was studied using a model system that generates free radicals (system CFL), culture of brain neurons, as well as models of stress in experimental animals.

The antioxidant properties of the potassium salt Kamenovo acid

The action of comenta potassium in the generation of reactive oxygen species (ROS) in the model system studied in the environment CFL (citrate-phosphate buffer with the addition of lyuminola) of the following composition: 4 ml citrate - phosphate buffer (105 mm KCl, 20 mm KH2PO4, 4 mm sodium citrate; pH was 7.45) with the addition of lyuminola (10 mm). The formation of ROS initiated introduction the m with constant stirring 30 μl of 35 mm solution of sulphate of iron. In this model, the oxidation of iron ions in the presence of orthophosphate and citrate accompanied by the formation of ROS and when this occurs chemiluminescence (CHL), selectively increasing lyuminola, which is suppressed in the presence of antioxidants. Registration chemiluminescence was carried out by the device SmartLum 5773 within 5 minutes. Estimated sutasoma chemiluminescence.

The antioxidative activity of the potassium salt Kamenovo acid was estimated by the inhibition of CHL a model system by adding aqueous solutions of the drug in comparison with the solution Kamenovo acid. The final concentration in the cuvette was 0.1 mg/ml and 0.01 mg/ml Chemiluminescence free radicals model system CPL (control) is taken as 100%. The results of the experiments were determined by the intensity of chemiluminescence (USD) and was calculated as a percentage of control [Farkhutdinov P.P., Lihovsky VA Chemiluminescent methods for the study of free radical oxidation in biology and medicine. 1995, Ufa, 90 S.]. The data processing was performed using software PowerGraph version 3.3. Reliability of differences was assessed using student's criterion. The results of the study are presented in table 1.

Table 1
The reduction of free radicals in the model system CFL in the presence of Kamenovo acid and its potassium salt (% of control)
№ p/pMedicationConcentration, mg/ml
0,010.1
1coment potassium34,24±1,22*64,6±0,95***
2kamenova acid33,29±1,36*at 69,39±1,16***
Note:
1*p<0.001 in comparison with control;
2**p<0.001 in comparison with 0.01 mg/kg

Analysis of the data presented in table 1 shows that the potassium salt of Kamenovo acid significantly reduces the content of free radicals in comparison with the control in the model system CFL. The level of reduction of free radicals by comentem potassium practically does not differ from the level of the quenching of free radicals Kamenovo acid. The antioxidant properties of comenta potassium and Kamenovo acid depends on the concentration of the substance. So, the increase of their concentration in the test process is e with 0.01 mg/ml to 0.1 mg/ml significantly (more than 30%) to increase the level of quenching free radicals.

Thus, the potassium salt of Kamenovo acid has strong antioxidant effect, while its antioxidant properties do not differ from those of Kamenovo acid.

For assessment of oxidative mechanisms in the brain most often used definition of products of lipid peroxidation (LPO), an intermediate product which is malonic dialdehyde (MDA), as well as the study of chemiluminescence biological environments.

The influence of comenta potassium on the intensity of free radical oxidation and MDA content in the brain of stressed animals.

The level of free radical oxidation in the brain was determined by chemiluminescent method [Farkhutdinov OR, Farkhutdinov P.P. // bul. the experts. Biol. and the honey. 2000, C. No. 3, P.260-264] on the device SmartLum 5773. The experimental results were evaluated according to the intensity of chemiluminescence (sutasoma chemiluminescence in USD) and was calculated as a percentage of control. Statistical analysis of results was performed using student's criterion [Lakin GF Biometrics. M. 1990. 352 C.]. MDA content was determined by the method Gavrilova BV et al. [Gavrilov V.B. have been, Gavrilova, A.R., Mazhul L.M. Determination of peroxide oxidation of lipids in the test with thiobarbiturate acid // Questions of medical chemistry. 1987. No. 1. P.19-21]. Statisticheskiy results was performed using student's criterion [Lakin GF Biometrics. M. 1990. 352 p] MO Excel 2003.

Coment potassium was administered to mice per os for 3 days, once a day on an empty stomach, at concentrations of 1, 2, 4 and 8 mg per 1 kg of body weight to stressaway mice. Stress in mice caused by hanging for cervical fold for five hours. After 5 hours the mice were decapotable, the brain was placed in liquid nitrogen and then in the brain was determined by the level of free radical oxidation (CPO) and MDA.

The interval calculation of quantities of comenta potassium 1-8 mg/kg of body weight was performed with regard to its effectiveness and lack of harmful effects on the body.

It is known that the toxic dose of potassium for humans is 6 g [Rock AV, Rudakov I.A. Bioelements in medicine. M. 2004. 272 C.]. The maximum single dose of potassium contained in nominate potassium per person in our experiments is 0.11, we have Also compared the content of K+our drug and medicines KCl and potassium Asparaginate. So, a single dose of potassium for humans in KCl is 0.21 g, and the medicines of potassium Asparaginate single dose of 0.04 g and daytime 0,12, In this regard, the use of comenta potassium in quantities of more than 8 mg/kg of body weight is considered inappropriate.

There were formed the following groups of male outbred mice:

1) intact;

2) intact + coment potassium 1 mg/kg;

3) intact + coment potassium 2 mg/kg of body weight;

4) intact + coment potassium 4 mg/kg;

5) intact + coment potassium 8 mg/kg of body weight;

6) tressirovanye;

7) tressirovanye + coment potassium 1 mg/kg;

8) tressirovanye + coment potassium 2 mg/kg of body weight;

9) tressirovanye + coment potassium 4 mg/kg;

10) tressirovanye + coment potassium 8 mg/kg of body weight. The experiment was conducted on 54 outbred mice-males, weighing 23-25 g, 9 mice in each group. The results obtained are presented in table 2.

Table 2
The influence of comenta potassium SRO and the content of MDA in the brain (trained mice (M±M)
№ p/pGroups of animalsSRO (sutasoma, $)MDA (nm/1 g protein)
1the intact208,23±or 4.313,17±0,14
2intact + coment potassium 1 mg/kg214,32±2,173,37±0,52
3intact + coment potassium 2 mg/kg 215,89±1,253,69±0,48
4intact + coment potassium 4 mg/kg221,12±4,913,32±2,17
5intact + coment potassium 8 mg/kg218,01±4.26 deaths3,49±1,23
6tressirovanye247,08±7,71**4,46±0,20***
7tressirovanye + coment potassium 1 mg/kg234,03±6,523,42±1,07
8tressirovanye + coment potassium 2 mg/kg228,13±5,12▪3,31±0,31▪
9tressirovanye + coment potassium 4 mg/kg218,00±3,62▪▪3,28±0,28▪
10tressirovanye + coment potassium 8 mg/kg204,99±2,27▪▪3,21±0,15▪▪▪
Note:*p≤0,05;**p≤0,01;***p≤0,001 - dostov most statistically significant differences compared with intact control
▪ p≤0,05; ▪▪ p≤0,01; ▪▪▪ p≤0,001 - reliability compared to tressirovanye animals (stress control).

Analysis of the data presented in table 2, shows that the application of comenta potassium in amounts 1-8 mg/kg of body weight in intact animals (2, 3, 4 5 group) level of free radical oxidation in the brain is virtually indistinguishable from the intact control (group 1). The sharp increase in the level of free radicals (p≤0,01) was observed after exposure to stress (group 6) on the body of animals. In the application of comenta potassium in amounts 1-8 mg/kg of body weight, the intensity of the SRO in the brain of stressed animals is reduced. In group 7 (1 mg/kg body weight) there is quite a weak reducing SRO. The most pronounced antioxidant activity of the potassium salt of Kamenovo acid is in an amount of 8 mg/kg body weight (group 10).

Investigation of the influence of comenta potassium content of MDA in the brain of the aforementioned groups of animals showed that coment potassium in amounts 1-8 mg/kg body weight practically no effect on MDA content in the brain of intact animals (groups 2, 3, 4, 5). Significant and reliable, in comparison with intact animals, the increase in MDA level observed after exposure to stress on the body (group 6). While in mice stressed and the floor is ausich coment potassium in quantities of 1, 2, 4 and 8 mg/kg of body weight has decreased MDA content in comparison with group 6 (stress control). The most pronounced and almost the same decrease in MDA level observed in groups of mice 8, 9 and 10 (2, 4 and 8 mg/kg body weight).

Thus, it was established that coment potassium in quantities of 2, 4 and 8 mg/kg of body weight has expressed an antioxidant stress-protective effect, resulting in intense decrease of hyperproductive free radicals and contents of the products of lipid peroxidation in the brain of stressed animals. The most pronounced antioxidant, stress-protective action of the potassium salt of Kamenovo acid assists in the amount of 8 mg/kg of body weight. In this regard for further research we have used potassium salt of Kamenovo acid number of 8 mg/kg of body weight.

The influence of the potassium salt Kamenovo acid on the antioxidant glutathione system in the brain of stressed animals

It is known that the antioxidant system is required to monitor the production of active forms of oxygen and prevent free radical reactions are enzymes, and numerous low-molecular antioxidants or compounds that prevent the formation of free radicals. Various antioxidants play in tissues different role.

In the author link in the protection of cells against the toxic effects of primary products FLOOR is the antioxidant glutathione system [Baraboi, VA, Sutkovoi D.A. Oxidative-antioxidant homeostasis in health and disease / Edited. Ed. by Acad. The Academy of medical Sciences of Ukraine Y.A. of zozula. K., 1997, 413 S.]. The influence of the potassium salt Kamenovo acid on the antioxidant glutathione system was assessed by the activity of the enzymes glutathione peroxidase, glutathione reductase and the content of reduced glutathione in the brain of stressed animals.

Glutathione performs the function of the donor hydrogen and cofactor of several antioxidant enzyme systems. The decrease in the intracellular content of reduced glutathione significantly reduces the resistance of cells and organism to intoxication. Determination of free sulfhydryl groups (reduced glutathione (GSH)in acid-soluble nadeshiko was determined by the method of [Sedlak j, Lindsay R.H. Estimation of total, proteinbound and nonprotein sulthydryl groups in tissue with Ellman''s reagent // Anal. Biochem. 1968. Vol.25. P.192-205].

The enzyme glutathione peroxidase (GP) is one of the main antioxidant enzymes protect the body from endogenous or exogenous-induced formation of peroxides, including lipid peroxides. Using glutathione SE restores the main product of lipid peroxidation - gidroperekisi lipids, neutralizing their toxic effects on the membrane and prevents the initiation of secondary oxidation reactions of lipids [Gavrilova, A.R., Khmara NF Determination act is vnesti mutationprobability of red blood cells with saturating concentrations of substrate // Laboratory work. 1986. No. 12. S-724].

The glutathione peroxidase was determined by the rate of oxidation of glutathione restored in the presence of gidroperekisi tertiary butyl [Moin V.M. Simple and specific method for determining the activity of glutathione peroxidase in erythrocytes // Laboratory work. 1986. No. 12. S-727]. The essence of the method lies in the fact that during incubation, samples of the glutathione peroxidase from the supernatant oxidizes saturating concentrations of glutathione restored substrate. The number of remaining non-oxidized glutathione restored judged on the activity of glutathione peroxidase.

The functional role of high-affinity cytoplasmic glutathione reductase is the generation of reduced glutathione from its disulfide form.

The activity of glutathione reductase (GR) is determined according to the method of [Yusupov LB ON improving the accuracy of determining the activity of erythrocyte glutathione peroxidase // Laboratory work. 1989. No. 4. P.100-101].

The experiment was carried out on outbred mice - males, weighing 18-20 g Were formed the following groups of animals:

1) intact;

2) tressirovanye;

3) intact + coment potassium 8 mg/kg of body weight;

4) tressirovanye + coment potassium 8 mg/kg of body weight.

Coment potassium was administered to mice at a concentration of 8 mg/kg body weight for 3 days before stressaway belly is s, once a day on an empty stomach. The stress caused by the method of immobilization by hanging mice for cervical fold for five hours. After 5 hours the mice were decapotable, the brain was placed in liquid nitrogen and then determined the content of reduced glutathione, the activity of the enzymes glutathione peroxidase and glutathione reductase.

The results of the study are presented in table 3.

Table 3
The influence of comenta potassium antioxidant glutathione system brain stressed animals
№ p/pGroups of animalsGSH µmol / 1G owner. TCSE umol / 1 min 1 g proteinG umol / 1 sec 1 g protein
1the intact2,00±0,05664,00±12,001,28±0,06
2tressirovanye1,82±0,04*745,00±30,00*1,47±0,05*
3intact + K is menat potassium 8 mg/kg of body weight 2,07±0,06▪▪612±24,00▪▪1,18±0,16
4tressirovanye + coment potassium 8 mg/kg of body weight2,04±0,08▪619,00±22,00▪▪1,24,±0,10▪
Note:
*p≤0,05;**p≤0,01 - reliability statistically significant differences compared with intact control;
▪ p≤0,05; ▪▪ p≤0,01 compared with tressirovanye animals (stress control).

Analysis of the data presented in table 3 showed that after exposure to stress (group 2) in the brain of animals significantly increased the activity of glutathione peroxidase (p≤0,001), increases the activity of glutathione reductase (p≤0,05), there has been a significant decrease in the concentration of reduced glutathione (p≤0,05).

The use of comenta potassium in an amount of 8 mg/kg body weight for 3 days before exposure to stress contributes to the normalization of antioxidant processes in the brain of experimental animals (gr. 4). Practically did not change indicators glutathione metabolism after administration of comenta potassium intact animals. That is, coment potassium in an amount of 8 mg/kg body weight has a neg is a high influence on glutathione metabolism in the intact animal. Thus, it was found that immobilization stress in mice leads to intensification FLOOR (high content of SRO and MDA) in the brain and mobilization of glutathione AOC (a statistically significant increase in the activity of the enzymes glutathione peroxidase and glutathione reductase). The use of the potassium salt Kamenovo acid number of 8 mg/kg of body weight for three days before stressaway animals contributes to the normalization of oxidative mechanisms FLOOR and, accordingly, the restoration of the antioxidant defense of the brain, which results in decreased activity of the enzymes glutathione peroxidase and glutathione reductase, increased concentrations of reduced glutathione. The level of antioxidant protection in the application of comenta potassium practically does not differ from that in the brain of intact animals (group intact control).

The influence of the potassium salt Kamenovo acid on glutamate cytotoxicity in culture granular cells of the cerebellum of rats

Development of methods of prevention and treatment is based on the assumption that a significant number of brain cells can be rescued by blocking the initiation or progress vnutrineironalnah pathogenicity destructive processes that have specific temporal and spatial characteristics.

Influence of potassium salt Komen is howling acid on glutamate cytotoxicity in culture neurons of the cerebellum of rats was studied in comparison with Kamenovo acid.

Studies were performed on Wistar rats. Culture of granular cells of the cerebellum were obtained from brain 7-9-day-old rat by the method of enzymatic and mechanical dissociation [Victorov IV, khaspekov L.G., Shashkova N.A. // guidance on the cultivation of nervous tissue: Methods, techniques, problems. 1986. M 266 C.]. Cell culture after 7 days of cultivation were subjected to the action of glutamate and/or potassium salts of Kamenovo acid, Kamenovo acid.

Coment potassium and Kamenovo acid made in group cultures of neurons without affecting glutamate and after a 10-minute exposure (100 μm) of glutamate in a balanced salt solution (SSR), pre-placing them in the original culture medium. Then the culture was incubated in CO2-the incubator for another 4.5-5 hours. The influence of comenta potassium in concentrations of 10-2M, 10-3M, 10-4M, 10-5M, 10-6M and Kamenovo acid in concentrations of 10-3M, 10-4M served as Control culture placed for 10 min in SSR, and cultures treated with glutamate in the SSR. After fixation, the cultures took into account the number of live and dead neurons on the inverted microscope Invertoscopes ID 03. Reliability of differences was assessed using student's criterion in MO Excel 2003.

Culture cerebellar neurons derived from intact rat was divided into the following groups, respectively

1.1 control SSR;

1.2 SSR + coment potassium 10-2M;

1.3 SSR + coment potassium 10-3M;

1.4 SSR + coment potassium 10-4M;

1.5 SSR + coment potassium 10-5M;

1.6 SSR + coment potassium 10-6M;

1.7 SSR + kamenova acid 10-3M;

1.8 SSR + kamenova acid 10-4M;

2.1 control SSR with glutamate (Glu) 100 µm;

2.2 SSR with glutamate (Glu) 100 µm + coment potassium 10-2M;

2.3 SSR with glutamate (Glu) 100 µm + coment potassium 10-3M;

2.4 SSR with glutamate (Glu) 100 µm + coment potassium 10-4M;

2.5 SSR with glutamate (Glu) 100 µm + coment potassium 10-5M

2.6 SSR with glutamate (Glu) 100 µm + coment potassium 10-6M;

2.7 SSR with glutamate (Glu) 100 µm + kamenova acid 10-3M;

2.8 SSR with glutamate (Glu) 100 µm + kamenova acid 10-4M. the Results of the study are presented in table 4.

Table 4 - Influence of comenta potassium and Kamenovo acid on the cytotoxic effect of glutamate in the culture of neurons of the cerebellum
№ p/pCropsThe number of live neurons (%)
ControlComent potassium (M) Kamenova acid (M)
10-210-310-410-510-610-310-4
12345678
1SSR95,12±0,6294,82±1,0595,13±0,9796,14±0,7695,92±0,7195,60±1,11to 85.8±3,3to 85.2±6.0
2glutamate29,04±1,56#**48,28±2,14#**46,88±1,35#**64,97±3,27#**54,65±3,51#**50,37±2,37#**46,7±3,1#**35,9±2,9#*
Note:
1#p<0,001 - in comparison with control SSR;
2*p<0,05 in comparison with the control group glutamate;
3**p<0,001 - in comparison with the control group glutamate.

Analysis of the results showed (table 4)that the number of surviving neurons after exposure of comenta potassium (group 1.2-1.6) and Kamenovo acid (group 1.7, 1.8) on intact granular cells of the cerebellum, regardless of the concentration of the substance, practically does not differ from the control group (1.1). After exposure to glutamate survival of granular cells of the cerebellum significantly (66,08%) reduced (group 2.1), p<0,001. Introduction in culture of cerebellar neurons treated with glutamate of comenta potassium (10-2M - 10-6M), contributed to significant increase in the resistance of neurons to glutamate cytotoxicity in all groups of crops (group 2.2-2.6). The maximum survival of neurons was observed in the group of crops 2.4 with the introduction of comenta potassium concentration of 10-4M When applying Kamenovo acid survival of neurons in cultures treated with glutamate (2.7 and 2.8)was significantly higher than that in the control group-glutamate, however, its maximum was significantly lower than when added to culture the potassium salt Kamenovo acid.

Thus, it was established that the use of the potassium salt Kamenovo acid in the face of glutamate cytotoxicity contributes to a significant increase in the survival of cerebellar neurons in culture. The sustainability of the neurons to the toxic effects of glutamate in cultures with the addition of the salt Kamenovo acid exceeds that in the application of Kamenovo acid, is used as a prototype for the greatest number of matches.

The toxicity of the potassium salt Kamenovo acid we were assessed by free radical oxidation and lipid peroxidation in the brain of experimental animals (table 2), as well as in the control cultures of cerebellar neurons with the addition of the investigated doses above salts. The results showed that the potassium salt of Kamenovo acid in the investigated doses not toxic.

It is known that the basis for clinical trials of antioxidants in the treatment of diseases and degenerative disorders are studies conducted in cultured cells and animal models [Andersen Julie K. Oxidative stress in neurodegene exploration: cause or consequence? // Nature Reviews Neuroscience 2004. Vol.5. S 18-S25].

We investigated the effect of different doses of potassium salts of Kamenovo acid on the levels of free radicals in the model system CFL and in the brain of stressed animals, on the antioxidant glutathione system in the brain (trained animals (table 1, 2, 3)and on the stability of cultured cerebellar neurons to glutamate cytotoxicity (table 4).

Polucheniyami evidence suggests, that stress on the organism of experimental animals was accompanied by increased oxidative processes in the brain, manifested in increased levels of free radicals and increased content in the brain of one of the products of lipid peroxidation, MDA, reduced glutathione protection.

It is established that the application of the potassium salt Kamenovo acid in quantities of 2 to 8 mg/kg of body weight to stress on the body significantly reduces the levels of free radicals, stimulates the antioxidant glutathione system in the brain of stressed animals. In these doses potassium salt of Kamenovo acid by oral administration has expressed an antioxidant, stress-protective action.

We studied the influence of the potassium salt Kamenovo acid on the stability of cultured cerebellar neurons to glutamate cytotoxicity.

It is established that the application of comenta potassium in concentrations of 10-2M - 10-6M in the culture of nerve cells of the cerebellum on the background of glutamate cytotoxicity significantly increase neuronal survival. The maximum survival of neurons observed in the group of cultures after exposure of comenta potassium concentration of 10-4M (group 2.9, p<0,001).

Getting comenta potassium and study by the authors of the subtle elements of the mechanism without istia on activated oxidative processes in the brain of stressed animals and the development of glutamate excitotoxicity leads to the conclusion about the prospects of its use as an antioxidant, stress and neuroprotective drugs in ischaemic brain damage, including violation of ion homeostasis and on the background of cardiovascular disease. Analysis of the known prior art did not allow to find a solution that exactly matches the set of essential characteristics with declare that confirms the novelty of the compounds as antioxidant, stress and neuroprotective funds. Thereby expanding range of highly effective drugs with antioxidant, stress and neuroprotective properties for the prevention and treatment of neurodegenerative diseases caused by oxidative damage to the brain.

Therefore, the claimed technical solution meets the criteria: novelty, inventive step and industrial applicability.

The use of the potassium salt Kamenovo acid as a prophylactic and therapeutic antioxidant, stress and neuroprotective agents in quantities of from 2 to 8 mg per 1 kg of body weight, fasting once in the course of 3 days.



 

Same patents:

FIELD: chemistry.

SUBSTANCE: invention relates to novel pyranyl aryl methylbenzoquinazolinone compounds of formula (I), which are positive allosteric modulators of the M1 receptor and which can be used to treat diseases associated with the M1 receptor, such as Alzheimer's disease, schizophrenia, pain disorders or sleep disturbance. In formula (I) X-Y are selected from a group comprising (1) -O-CRARB-, (2) -CRARB-O-, (3) -CRARB-SRC-, (4) -CRARB-NRC- and (5) -NRC-CRARB-, where each RA and RB is a hydrogen atom, and RC is selected from a group comprising (a) hydrogen, (b) -C(=O)-C1-6alkyl, (c) -C1-6alkyl, (d) -C(=O)-CH2-C6H5, (e) -S(=O)2-C1-6 alkyl, R1 is a hydroxy group, R2 is selected from a group comprising (1) -phenyl, (2) - heteroaryl, where the phenyl or heteroaryl group R2 is optionally substituted; the rest of the values of the radicals are given in the claim.

EFFECT: obtaining novel pyranyl aryl methylbenzoquinazolinone compounds.

28 cl, 12 tbl, 37 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine and pharmacology and concerns using comenic acid sodium salt as a preventive and therapeutic antioxidant, stress and neuroprotective agent in the amount of 1 to 4 mg per 1 kg of body weight daily on the empty stomach for 3 days.

EFFECT: invention provides the high clinical effectiveness.

3 tbl

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine, particularly to pharmacology and pharmaceutics, and concerns a sedative agent representing glycine immobilised on detonation nanodiamond particles 2-10 nm in size, and to a method for preparing it.

EFFECT: preparing the sedative agent.

4 cl, 7 dwg, 13 tbl, 4 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention discloses crystalline form of S-zopicone with spectrum of powder X-ray diffraction with application of Cu-Ka radiation, with characteristic peaks, expressed under conditions 20 approximately at 11.08°, approximately 12.38°, approximately 15.86°, approximately 17.88°, approximately 19.98°C and approximately 20.58°, DSC-thermogram, on which peak is observed approximately at 207.7°C, and infrared spectrum of absorption (IR) with characteristic peaks approximately at 3078 cm-1, approximately 2942-2838 cm-1, approximately 2790 cm-1, approximately 1716 cm-1, approximately 1463 cm-1, approximately 1372 cm-1 and approximately 757 cm-1.

EFFECT: claimed are: method of preparation of crystalline form of eszopiclone, pharmaceutical preparation and its application in manufacturing medication for treatment of sleep disorder.

8 cl, 8 dwg, 10 ex

FIELD: medicine.

SUBSTANCE: present group of inventions refers to medicine, namely therapy and neurology, and concerns a melatonin agonist therapy. As the above melatonin agonist, (lR-Trans)-N-[[2-(2,3-dihydro-4-benzofuranyl)cyclopropyl]methyl]propanamide is administered in effective doses that provides treating or preventing disruption of circadian rhythm or sleep disturbance.

EFFECT: invention provides treating or preventing disruption of circadian rhythm or sleep disturbance.

9 cl, 2 ex, 4 tbl

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention provides new imidazo[1,2-b]pyridazine compounds covered by general structural formula (I) wherein the radicals and symbols have the values presented in the patent claim, and pharmaceutically acceptable salts thereof. The compounds of structural formula (I) are effective both for treating or preventing the diseases related to GABA receptor inhibition, anxiety, epilepsy, sleep disorders, including insomnia, and for inducing a sedative-hypnotic, anaesthetic effect, sleep and muscle relaxation.

EFFECT: there are presented methods for preparing the above compounds, and also intermediate compounds for preparing them.

21 cl, 4 tbl, 13 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine and pharmaceutical industry, and concerns a pharmaceutical composition for sublingual or buccal administration of the active ingredients of a low to poor water-solubility. As an active ingredient, the composition contains a solution of a hormone specified in a group consisting of melatonin, oestrogens, progesterone, testosterone and dihydrotestosterone in a pharmaceutically acceptable solvent, adsorbed or absorbed on particles of a pharmaceutically acceptable carrier. The invention also concerns methods for preparing and using the above pharmaceutical composition.

EFFECT: what is presented is the new composition for sublingual or buccal administration of the active ingredients of a low to poor water-solubility.

19 cl, 4 ex, 2 tbl, 2 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: what is presented is the use of 7-chlor-3-(5-dimethylaminomethyl-[1,2,4]oxadiazol-3-yl)-5-methyl-4,5-dihydro-imidazo[1,5-a][1,4]benzodiazepin-6-one or a pharmaceutically acceptable salt thereof for treating various types of insomnia (terminal one, insomnia in an individual under min. 65 years, for the relief of wakefulness after the beginning of a sleep, for the prolongation of total sleep time after the beginning of a sleep - versions), pharmaceutical compositions for the appropriate application (versions) and methods of treating the various types of insomnia (versions).

EFFECT: invention is effective in treating insomnia, maintaining the sleep, terminal insomnia, including in the aged; the compound has a short half-life, ie 3-4 hours (so it causes no residual sedative action), and improves the daily activity in the aged suffering diurnal drowsiness.

61 cl, 29 dwg, 10 tbl

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to compounds of formula (I) and salts thereof wherein R1 represents -A11-A12-; R2 represents tetrahydrofurylmethyl, tetrahydropyranylmethyl or tetrahydropyranyl; A11 represents a single bond, methylene or 3,2-ethylene; A12 represents C1-6 alkyl, C3-6 cycloalkyl or C3-6 cycloalkyl containing methyl; R3 represents methoxy, cyano, cyclobutyloxymethyl, methoxymethyl or ethoxymethyl; and R4 represents methoxy or chlorine. Also, the invention also refers to a pharmaceutical composition possessing corticotrophin-releasing factor (CRF) receptor antagonist activity, containing a compound of formula (I), to a therapeutic/preventive agent, and a method of treating the diseases specified in the patent claim.

EFFECT: there are presented the compounds of formula (I) as corticotropin-releasing factor (CRF) receptor antagonists.

20 cl, 2 dwg, 2 tbl, 51 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to new halogenised pyrazolo[1,5-a]-pyrimidines of general formula (I) and their pharmaceutically acceptable salts possessing affinity with respect to α1-,α2 subunits of a GABAA receptor. In formula R represents alkyl(C1-C6); R1 is specified in a group consisting of alkyl(C1-C6) and alkinyl(C1-C6); X represents a halogen atom, and Y is specified in a group consisting of -CO- and -SO2. The invention refers to intermediate enamine compounds and methods for preparing them.

EFFECT: invention also refers to a method for preparing the compounds of formula (I), the based pharmaceutical compounds, to the use of said compounds for preparing said drug preparation for treating or preventing anxiety, epilepsy, sleep disorders, including insomnia, as well as for inducing a sedative-hypnotic effect, anaesthesia and muscular relaxation.

23 cl, 6 tbl, 10 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to a kit for lung cancer cell sensitisation to cisplatin. The declared kit comprises a first composition containing a therapeutically effective amount of sodium metaarsenite, and a second composition containing a therapeutically effective amount of cisplatin. Also, the invention refers to using the therapeutically effective amount of sodium metaarsenite for lung cancer cell sensitisation in a patient treated with cisplatin.

EFFECT: invention provides increasing the therapeutic effectiveness with a reduced risk of side effects.

10 cl, 4 tbl, 3 dwg, 1 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine and pharmacology and concerns using comenic acid sodium salt as a preventive and therapeutic antioxidant, stress and neuroprotective agent in the amount of 1 to 4 mg per 1 kg of body weight daily on the empty stomach for 3 days.

EFFECT: invention provides the high clinical effectiveness.

3 tbl

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely to rheumatology and may be used for treating the patients with ostheoarthrosis fulfilling driving and supervision functions with Theraflex Advance, Theraflex and Theraflex M cream. The treatment is two-staged, including 17-day hospital treatment and six-month outpatient treatment. The hospital treatment involves Theraflex Advance 2 tablets 3 times a day in a combination with local skin applications of Theraflex M cream 2-3 times a day as a 2-3 cm strip rubbed until absorbed completely. The outpatient treatment involves prescribing Theraflex 1 capsule 3 times a day for the first three weeks, further 1 capsule 2 times a day in a combination with Theraflex M cream locally applied on the skin 2-3 times a day as a 2-3 cm strip rubbed until absorbed completely.

EFFECT: invention provides the higher quality of life in the given category of patients ensured by the developed regimen of treatment enabling the complete withdrawal of myorelaxants and non-steroidal anti-inflammatory preparations.

1 tbl, 2 ex

FIELD: biotechnologies.

SUBSTANCE: invention relates to compounds of general formula I, having cytostatic or cytotoxic activity, its pharmaceutically acceptable salts, tautomers or stereoisomers, a pharmaceutical composition on their basis. Compounds may be used for treatment of cancerous diseases. In the general formula I

Y is selected from the group comprising -CHRay- and -CHRay-CRby-CRcy-; each Ray, Rby and Rcy are independently selected from hydrogen and non-substitute C1-C12-alkyl; each R1, R2, R3, R4 and R5 are independently selected from hydrogen and non-substitute C1-C12-alkyl; R6 is selected from NR8R9 and OR10; A means , W means NR7; R7 means hydrogen; R8 means hydrogen; R10 means non-substitute C2-C12-alkenyl; each dotted line means unnecessary additional link, but when there is a triple link between atoms of carbon, to which R1 and R2 are attached, then R1 and R2 are absent, and when there is a triple link between carbon atoms, to which R3 and R4 are attached, then R3 and R4 are absent; R9 is selected from substitute C2-C12-alkenyl and substitute C4-C|2-alkenylyl, where substitutes are selected from the group, consisting of: halogen, OR', OCONHR' and OH, protected with a simple silyl ether; where R' means hydrogen; provided that whenever Y means -CHRay-CRby=CRcy- and there is a single or double link between atoms of carbon, to which R3 and R4 are attached, then R9 means substitute C4-C12-alkenylyl; and each R16, R17 and R18 are independently selected from hydrogen and ORa; each Ra is selected from hydrogen or non-substitute C1-C12-alkyl.

EFFECT: higher efficiency of compound application.

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention refers to new 4-(heterocycloalkyl)benzene-1,3-diol compounds described by general formula (I) presented below, wherein: R1, R2, R3 and R4 may be identical or different, and represent: - hydrogen,- C1-C4-alkyl radical,- hydroxymethyl, hydroxyethyl, - (C1-C4-alkoxy) carbonyl,- C1-C4-alkoxy,- hydroxyl, or R1 and R2 are bound to each other and to a carbon atom whereto attached to form a carbon cycle containing 5 or 6 carbon atoms, while R3 and R4 may be identical or different, and represent: - hydrogen, - C1-C4-alkyl radical, or R1 and R4 are bound to each other to form the chain -(CH2)2- or -(CH2)3-, while R2 and R3 may be identical or different, and represent: - hydrogen, - C1-C4-alkyl radical, X represents an oxygen atom or a sulphur atom, Y represents hydrogen, a chlorine atom or a fluorine atom, the value of m may represent 1 or 2, while the value of n may represent 0 or 1, and provided n=0, then m=1 or 2, and provided n=1, then m=1, as well as to salts of the compounds of general formula (I) and to their isomer and enantiomer forms.

EFFECT: invention relates to the use of these compounds in pharmaceutical or cosmetic compositions for the use in treating or preventing pigmentary disorders.

7 cl, 2 tbl, 19 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to experimental medicine and pharmacology, namely a method for preparing lithium salt of comenic acid (lithium comenate) by mixing a comenic acid solution heated up to temperature max. 80±2°C, with lithium carbonate or hydroxide solution in the stoichiometrically-defined amounts, and colouring the solution in yellow and reducing pH of the solution to 4.6 or 10.0 respectively, and recovering luthium salt of comenic acid from the solution by water distillation under vacuum. For this purpose, lithium salt of comenic acid is suggested to be used in the amount of 2 mg per 1 kg of body weight once daily for 3 days.

EFFECT: invention refers to the agents for preventing and treating neurodegenerative diseases caused by an brain oxidative damage.

2 cl, 2 dwg, 5 tbl

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention refers to pharmacology and medicine, and concerns an antibactrial agent representing a nanodiamond conjugate to amikacin of particle size 2-10 nm with the amikacin content up to 40 wt %, as well as a method for preparing it.

EFFECT: preparing a new antibacterial agent.

3 cl, 6 dwg, 1 tbl, 1 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to a pharmaceutical combination and to its use for treating an infection caused by influenza virus. The declared composition contains a pyrazine derivative of formula wherein R1 and R2 are identical or different, and each represents a hydrogen atom or a halogen atom; and R3 represent a hydrogen atom or a protective group for amino group or its salt, and, and a neuraminidase inhibitor. The neuraminidase inhibitor is specified in oseltamivir, zanamivir, peramivir or CS-8958.

EFFECT: invention provides preparing the combination which shows strong antiviral activity, smaller side effects and applicable for treating influenza.

12 cl, 8 tbl, 4 ex

FIELD: chemistry.

SUBSTANCE: invention relates to a compound of formula or a pharmaceutically acceptable salt thereof, in which R1 denotes hydrogen or C1-6alkyl; R2 denotes isooxazolyl group, substituted with C1-6alkyl; RB denotes -CF3, -CHF2, -CH2F, or C1-6alkyl. The invention also relates to pharmaceutical compositions for treating cancer which contain the disclosed compounds.

EFFECT: obtaining novel compounds and a pharmaceutical compositions based on said compounds, which can be used in medicine for treating cancerous diseases.

15 cl, 77 dwg, 10 tbl, 13 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmaceutics and medicine and concerns the use of 3,5-dihydroxytoluene or its derivatives for preparing drugs and functional food stuff for treating or preventing depression.

EFFECT: developing the antidepressant representing 3,5-dihydroxytoluene or its derivatives showing more significant antidepressant activity as compared with fluoxetine or imipramine.

39 cl, 21 tbl, 9 ex

FIELD: pharmaceutical industry and technology, pharmacy.

SUBSTANCE: invention relates to the improved method for preparing dihydroquercitin. Invention involves preliminary separation of volatile nonpolar compounds with steam at rapid change of pressure from 0.1 to 0.3 MPa in reactor carried out before extraction stage and with reduced time of raw extraction being without milling wood under conditions of mild mechanical-chemical effect and without destruction of wood occurring in rapid change of pressure from 0.1 to 0.5 MPa using mixture of polar solvent with water or their mixtures as an extractant. Invention reduces extraction time in intensifying and optimization of the process of dihydroquercitin that provides reducing total time of process from 60-75 min to 7-9 min and to simplify technology in carrying out the process without milling wood.

EFFECT: improved method for preparing.

5 ex

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