Antioxidant, stress and neuroprotective pharmacological agent sodium comenate

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

 

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) [MS Popova, Induced M. Induction of cell cycle, amyloid-beta and free radicals in the mechanism of 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]. 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 in the result in the 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 [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. With 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; Zavalishin I.A., M. Zakharova. Multiple sclerosis: modern aspects of etiology and pathogenesis. // Journal of neurology and psychiatry. CSV (Mgr. "Multiple sclerosis"). 2003. No. 1. P.72-75].

Another important factor causing the death of neurons is prolonged stimulation of neurons in the Central nervous system neurotransmitter glutamate. As vozbuzhdaemym 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 and satiation re is atarov neighboring neurons and the damage zone extends Over-stimulation of glutamate receptors, 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 [Z.A. Suslina, 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. 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., N. Katsube ad Chuang 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. Saint-Petersburg. 2010. P.60].

Known use of sodium oxybutyrate (sodium salt of g-aximilian the th acid). Sodium oxybutyrate has a pronounced antihypoxic effect. It increases the stability of the body, including the tissues of brain, heart, retina, to oxygen deficiency. The drug has a sedative and Central miorelaksantnoe in high doses cause sleep and the state of anesthesia. The drug is contraindicated in hypokalemia, male. Caution is required when toxicosis of pregnant women with hypertensive syndrome. In connection with the sedative effect is not supposed to assign the product (nervousness, glaucoma etc) in the daytime persons whose work requires quick physical and mental reactions [http://ru.wikipedia.org/wiki; Kryzhanovsky GN. The possibility of using lithium oxybutyrate in pathology. // New data on the pharmacology and clinical use of lithium salts: proceedings of the Institute of pharmacology of the AMS SSR. Edited by Dr. med. Sciences, Professor Lyubimov. M. 1984. 49-57].

Known antioxidant neuroprotective agent pikamilon - sodium salt of N-nicotinoyl-γ-aminobutyric acid. The presence of the chemical structure of picamilone gammaaminobutyric acid (GABA) and nicotinic acid accounts for its nootropic and vasoactive effects. It improves blood circulation in the brain and has a positive effect on the metabolism of brain tissue. The drug has also antihypoxic and antioxidant antiagregantnoe action has a tranquilizing effect. The mechanism of action of picamilone due to its interaction with GABA-a receptors. Has side effects: allergic reactions, headache, dizziness, anxiety [Pharmacology of sport. / Gorchakov N.A., goudelock AS, Gunina L.M. and others; under the General Ed. S.A. Olenina, L.M. Gunina, RD of sejfulla. K.: Olympus. l-RA. 2010. 640].

The closest to saleimoa 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 S.].

Kamenova acid has a mild sedative effect, not causing addiction [RF patent for the invention №2209062, IPC7AC 31/351, OR 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 [Pat the NT of the Russian Federation No. 2322977, AC 31/351 (2006.01), OR 29/02 (2006.01)]. Has antiobscenity, anxiolytic and antidepressantlike 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 various neurodegenerative diseases associated with oxidative damage to 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, the sodium salt of Kamenovo acid (coment sodium)obtained by mixing solution Kamenovo acid, heated to a temperature of 80±2°C, with a solution of carbonate or sodium hydroxide in the amounts taken from the calculation of stoichiometric, until a pH value of 4.6. Coment sodium is used in an amount of from 1 to 4 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 sodium salt Kamenovo acid with sodium carbonate take 25 ml of 0.12 called hot solution is Kamenovo acid (80±2°C) and add 15 ml of 0.2 N. solution of sodium carbonate, stirring occasionally.

To obtain the sodium salt Kamenovo acid with sodium hydroxide take 25 ml of 0.12 called hot solution of Kamenovo acid (80±2°C) and add 30 ml of 0.1 n sodium hydroxide solution, stirring occasionally.

The reactions take place quickly, the solution turns yellow. Sodium 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 yield of sodium salt of Kamenovo acid is 85-90%.

It is known that sodium is important for nerve and muscle activity, since it forms an electrolytic environment (electrolytic environment is created in the main 3 elements - sodium, chlorine and potassium), essential for normal functioning of nerves and muscle cells [http://www.lishnemuvesu.ru/natrii.html]. Sodium is involved in the maintenance of osmotic pressure and acid-base balance, nerve impulses [http://vitameal.ru/ibod.php?h=mna].

It was also established that in addition to the regulation of osmosis, the generation of the action potential sodium plays a number of important functions in the Central nervous system.

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

D. the op perate comenta sodium to generate ROS in the model system studied in the environment CFL 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 by the introduction under stirring with 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 the 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 sodium 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 of Chemiluminescence free radicals model system CPL (control) is taken as 100% [Farkhutdinov P.P., Lihovsky VA Chemiluminescent methods of research of free-radical oxidation in biology and medicine. 1995. The Ufa. 90 C.]. The results of the experiments were determined by the intensity of chemiluminescence (USD) and was calculated as a percentage of control. The data processing was performed using software PowerGraph version 3.3. Reliability of differences was assessed using student's criterion. The results of the research the research presented in table 1.

Table 1
The reduction of free radicals in the model system CFL in the presence of Kamenovo acid and its sodium salt (% of control)
№ p/pMedicationConcentration, mg/ml
0,010,1
1Coment sodium29,90±1,27*67,72±1,20***
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 0.1 mg/ml solution of the sodium 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 sodium practically does not differ from the level of damping available for the Alov Kamenovo acid. The antioxidant properties of comenta sodium and Kamenovo acid depends on the concentration of the substance. Thus, increasing the concentration in the test solution with 0.01 mg/ml to 0.1 mg/ml significantly, by more than 30%, the increase of the level of quenching free radicals.

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

The influence of the sodium salt of Kamenovo acid on antioxidant 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. Superoxide dismutase (SOD) controls cellular levels of superoxide anion, dismutase its excess in the hydrogen peroxide. Catalase and glutathione enzymes turn the water hydroperoxide, if he does not leave the intracellular space. In the brain antioxidant enzyme system is represented mainly by superoxiddismutase and glutathione-dependent enzymes. [Fedorova so-CALLED. OK slitely stress and protect the brain from ischemic damage: Diss. Dr. Biol. Sciences: 03.00.04: M. 2004. 298 S. RSL OD 71:05-3/6]. SOD was determined by the method of E.E. of Dubinina [Dubinin E.E. and other Comparative analysis of the activity of superoxide dismutase and catalase erythrocytes and whole blood in infants with chronic hypoxia. Laboratory work 1988. No. 8. P.16-19].

Catalase activity was determined according to the method described M.A. Korolyuk and others [Method of determination of catalase activity. // Laboratory work. 1988. No. 1. P.16-19].

An important link in the protection of cells against the toxic effects of primary products of lipid peroxidation (LPO) is the antioxidant glutathione system [Baraboi, VA, sutkovoi D.A. Oxidative-antioxidant homeostasis in health and disease. / Under. Ed. by Acad. The Academy of medical Sciences of Ukraine Y.A. of zozula. K. 1997. 413]. The influence of the sodium salt of 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 over the CORF) was performed according to the method J.Sedlak [J. Sedlak, 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 the Determination of the activity of glutathione peroxidase in erythrocytes in 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 remaining number is not oxidized glutathione restored judged on the activity of glutathione peroxidase.

The functional role of highly specific cytoplasmatic the second glutathione reductase is the generation of reduced glutathione from its disulfide form.

The activity of glutathione reductase (GR) is determined according to the method LB Yusupov [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

Coment sodium was administered to mice per os, on an empty stomach, at doses of 1, 2 and 4 mg/kg body weight for 3 days, before stressaway animals once per day. 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, SOD and catalase. Dose of comenta sodium were determined on the basis of preliminary studies. The analysis of these experiments showed that coment sodium used in lower quantities, less than 1 mg/kg of body weight, has almost no influence on antioxidant processes in the brain of stressed animals. In the application of comenta sodium in amounts higher than 4 mg/kg of body weight, efficiency is not significantly different from data in the application in the amount of 4 mg/kg of body weight.

There were formed the following groups of animals:

1. intact;

2. intact +coment sodium 1 mg/kg of body weight;

3. intact + coment sodium 2 mg/kg of body weight;

4. intact + coment sodium 4 mg/kg body weight;

5. tressirovanye;

6. tressirovanye + coment sodium 1 mg/kg of body weight;

7. tressirovanye + coment sodium 2 mg/kg of body weight;

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

The results of the study are presented in table 2.

tressirovanye + coment sodium 2 mg/kg of body weight
Table 2
The influence of comenta sodium on the antioxidant system of the brain stressed animals
№ p/pGroups of animalsGSH mmol/1 g VL. clothSE µmol GSH/1 min 1 g proteinGR µmol NADPH/1 sec 1 g proteinSOD srvc. u/1 mg proteinCatalase µmol H2About2/min mg protein
1the intact1,99±0,03654,70±15,01,29±0,0429,00±2,430,576±0,07
2intact + coment sodium 1 mg/kg of body weight 1,93±0,06▪588,00±16,0▪▪▪1,29±0,08▪29,26±1,42▪0,628±0,33▪
3intact + coment sodium 2 mg/kg of body weight;1,93±0,06▪704,00±PHP 64.001,27±0,03▪▪27,33±1,460,632±0,065
4intact + coment sodium 4 mg/kg of body weight1,77±0,06604,0±33,0▪1,38±0,16of 28.27±1,940,628±0,024▪▪
5tressirovanye1,77±0,03***735,00±21,00*1,49±0,04**25,05±1,140,745±0,023*
6tressirovanye + coment sodium 1 mg/kg of body weight1,95±0,04▪▪624,00±16,0▪▪1,22±0,05▪▪29,76±1,89▪0,598±0,04▪▪
72,02±0,08▪▪617±38,001,30±0,1128,15±2,210,622±0,03▪▪
8tressirovanye + coment sodium 4 mg/kg of body weight1,86±0,04662±14,001,36±0,04▪27,12±1,39China 0,686±0,02
Note:
*p≤0,05;**p≤0,05;***p≤0,01 - reliability 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, showed that the introduction of intact mice per os sodium salt of Kamenovo acid in quantities of 1 and 2 mg/kg body weight (group 2, 3) the activity of the enzymes glutathione peroxidase, glutathione reductase and the content of reduced glutathione remained at the level of the intact control. Not detected changes in these groups and in the activity of the enzymes SOD and catalase. In the application of comenta sodium 4 mg/kg body weight (group 4) level restored gluta the ion decreased to the level of stressed animals (p> 0.05), and was slightly increased the enzymatic activity of glutathione reductase, did not differ from intact control the activity of glutathione peroxidase. No significant changes in the application of comenta sodium 4 mg/kg body weight (group 4) in the activity of the enzymes SOD and catalase.

Stress effects on the body of mice (group 5) is accompanied by pronounced changes in the antioxidant glutathione protection. Thus, increases in comparison with intact animals in the brain activity of glutathione peroxidase (p≤0.05), and increases the enzymatic activity of glutathione reductase (p≤0.01), and significant decrease in the concentration of reduced glutathione (p≤0,001).

With the introduction of comenta sodium per os in quantities of 1 and 2 and 4 mg/kg body weight (group 6, 7, 8) for three days before stressaway in the brain of these groups of animals celebrated the restoration of the antioxidant glutathione protection to its level in the intact animals.

Under stress in mice, a marked reduction of the enzymatic activity of SOD and the increase of catalase activity (group 5). In the application of comenta sodium in quantities of 1, 2 and 4 mg/kg of body weight, the activity of SOD and catalase in the brain of stressed mice almost recovered to the level of the intact animal group 6, 7, 8).

Thus, it was found that immobilization stress in mice leads to mobilization of glutathione AOC (a statistically significant increase in the activity of the enzymes glutathione peroxidase and glutathione reductase), the decrease of SOD activity and increased activity of catalase in the brain.

The sodium salt of Kamenovo acid in quantities from 1 to 4 mg/kg of body weight once a day for three days before stressaway animals contributes to the normalization of antioxidant protection for the brain. This is reflected in the decreased activity of the enzymes glutathione peroxidase and glutathione reductase, increased concentrations of reduced glutathione. It was also established that the use of comenta sodium in amounts of from 1 to 4 mg/kg of body weight promotes recovery from stress influence the enzymatic activity of SOD and catalase. The level of antioxidant protection in the application of comenta sodium practically does not differ from that in the brain of intact animals (group 1).

The influence of the sodium salt of 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 and and move vnutrineironalnah pathogenicity destructive processes, with certain temporal and spatial characteristics.

The influence of the sodium salt of Kamenovo 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]. Cell culture after 7 days of cultivation were subjected to the action of glutamate and/or comenta sodium, Kamenovo acid.

Coment sodium 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 sodium at 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 in inverte is consistent 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 sodium 10-2M;

1.3 SSR + coment sodium 10-3M;

1.4 SSR + coment sodium 10-4M;

1.5 SSR + coment sodium 10-5M;

1.6 SSR + coment sodium 10-6M;

1.7 SSR + kamenova acid 10-3M;

1.8 SSR + kamenova acid 10-4M;

1.9 control SSR with glutamate (Glu) 100 µm;

2.0 SSR with glutamate (Glu) 100 µm + coment sodium 10-2M;

2.1 SSR with glutamate (Glu) 100 µm + coment sodium 10-3M;

2.2 SSR with glutamate (Glu) 100 µm + coment sodium 10-4M;

2.3 SSR with glutamate (Glu) 100 µm + coment sodium 10-5M

2.4 SSR with glutamate (Glu) 100 µm + coment sodium 10-6M;

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

2.6 SSR with glutamate (Glu) 100 µm + kamenova acid 10-4M

The results of the study are presented in table 3.

Table 3
The influence of comenta sodium and Kamenovo acid on the cytotoxic effect of glutamate in the culture of neurons of the cerebellum
№ p/p CropsThe number of live neurons (%)
controlComent sodium (M)Kamenova acid (M)
12345678
10-210-310-410-510-610-310-4
1SSR95,12±0,6283,78±1,8784,53±1,9993,99±0,7693,81±0,7889,74±0,32to 85.8±3,3to 85.2±6,0
2glutamate29,04±1,56#**25,61±6,1242,69±2,54#** 52,34±2,35#**51,27±3,81#**26,94±5,0946,7±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.

The analysis of the results reflected in table 3, showed that the number of surviving neurons after exposure of comentou sodium (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. Almost at the same level and remains neuronal survival when added to cultures with glutamate of comenta sodium at a concentration of 10-2M and 10-6M (group 2.2 and 2.6). Improving the sustainability of cerebellar neurons to glutamate cytotoxicity is observed with the introduction in the culture treated with glutamate of comenta sodium at concentrations of 10-3M-10-5M (group 2.3 and 2.4). When applying Kamenovo acid neuronal survival in groups Kul is ur 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 the culture of the sodium salt of Kamenovo acid.

Thus, it was found that the sodium salt of 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 sodium salt Kamenovo acid exceeds that in the application of Kamenovo acid.

The toxicity of the sodium salt of Kamenovo acid we were assessed by antioxidant defense 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-mentioned salts. The results showed that the sodium salt of Kamenovo acid in quantities of 1-4 mg/kg of body weight is 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. S18-S25].

We investigated the effect of different doses of sodium salt to the ene acid on free radical oxidation in vitro model system CFL (table 1), antioxidant system in the brain of stressed animals (table 2), as well as its impact on the survival of cultured cerebellar neurons against the background of glutamate cytotoxicity.

The results of researches of influence of comenta sodium on free radical oxidation in vitro model system CFL) and antioxidant system brain in vivo suggest that the sodium salt of Kamenovo acid has antioxidant properties. The use of comenta sodium in quantities of 1-4 mg/kg of body weight, once daily for 3 days before stressaway mice contributes to the restoration of the antioxidant system of the brain. The most pronounced antioxidant, stress-protective effect with oral administration of the sodium salt of Kamenovo acid provides at 2 mg/kg of body weight.

It was also established that the use of the sodium salt Kamenovo acid in the culture of cerebellar neurons against the background of glutamate cytotoxicity significantly increases the stability of cerebellar neurons to glutamate cytotoxicity. The maximum survival of neurons in the background glutamate cytotoxicity observed in the group of cultures after exposure of comenta sodium at a concentration of 10-4M, 10-5M (group 2.4, 2.5).

It is known that ion homeostasis is a key biochemically the indicator organism in norm and pathology, and one of the most important and attractive differentiated targets for pharmacological interventions. In a series of macro - and micronutrients determine the level of health and a number of important physiological responses, including the nervous system, essential belongs to the sodium.

Earlier it was mentioned that sodium is important for nerve and muscle activity, since it forms an electrolytic environment, it is necessary to maintain osmotic pressure and acid-base balance, for the normal functioning of nerves and muscle cells in the conduction of nerve impulses [Rock AV, Rudakov I.A. Bioelements in medicine. M. 2004. 272 C.]. Sodium plays a number of important functions in the CNS. So the transporters of glutamate based on the gradient of sodium ions, potassium and amino acids [Semenov AV, Kazantsev V.B. have been the Neuron-glial interactions in the brain. Nizhny Novgorod. 2007. 107]. From sodium dependent transport of amino acids, sugars, various inorganic and organic anions across the cell membrane. For a man of sodium is non-toxic.

Getting comenta sodium and study by the inventors of the subtle elements of the mechanism of its effect on activated oxidative processes in the brain of stressed animals and the development of glutamate excitotoxicity, p is help make a conclusion about the prospects of its use as an antioxidant, stress and neuroprotective agents in neurodegenerative diseases, including those associated with impaired ion homeostasis. Analysis of the known prior art confirms the novelty of the compounds as antioxidant, stress and neuroprotective agents.

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 sodium salt of Kamenovo acid as a prophylactic and therapeutic antioxidant, stress and neuroprotective agents in amounts of from 1 to 4 mg per 1 kg of body weight, fasting once in 3 days.



 

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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 pharmaceutical composition for treating and/or preventing depressions. The pharmaceutical composition contains an active substance presented by a selective serotonin reuptake inhibitor (SSRI) specified in a group of fluoxetine, paroxetine, citalopram, escitalopram, sertraline, fluvoxamine differing by the fact that as an active substance, it additionally contains N-acetyl-5-methoxytryptamine (melatonin) in the following proportions, mg: selective serotonin reuptake inhibitor (SSRI) - 10-30 mg, melatonin - 3-8 mg. The pharmaceutical composition may be presented by a solid dosage form - a tablet, a film-coated tablet, a capsule, by a soft dosage form - a rectal suppository.

EFFECT: pharmaceutical composition provides treating depressions and has a number of additional therapeutic properties: easing falling asleep and relieving sleeping disorders, recovering circadian rhythm and seasonal rhythm with reducing a risk of side effects of SSRI.

3 cl, 14 tbl, 7 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention refers to a new (-)-stereoisomer of formula (I) wherein X is H, or its pharmaceutically acceptable salt which agonise GABA receptor, to a pharmaceutical composition on the basis of the presented compound, to a method for preparing the (-)-stereoisomer of formula (I) or its pharmaceutically acceptable salt, to a method for inducing or maintaining general anaesthesia, to a method for promoting pain management and to a method for promoting pain management and to a method for prototyping antiemetic activity with the use of the presented (-)-stereoisomer or its pharmaceutically acceptable salt, as well as to a new diastereoisomer (-)-2,6-di-fluoro-butylphenyl ester of carbamic acid of formula (II) wherein R1 represents a chiral amino group, and X is H, or to its pharmaceutically acceptable salt.

EFFECT: preparing the pharmaceutically acceptable salt which agonise GABA receptor.

14 cl, 15 ex, 8 tbl, 3 dwg

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: medicine, pharmaceutics.

SUBSTANCE: present invention refers to two-layer tablet containing a compartment comprising a pharmaceutically acceptable compound of simvastatin; and a simultaneously separate compartment containing a compound of lisinopril and a pharmaceutically acceptable compound of folic acid as active ingredients. Preferentially, the tablet is prepared by using either direct compression technique, or wet granulation technique. The two-layer tablet under the invention is applied for purposes of preventing cardiovascular disturbances, including apoplexy and paralysis accompanying diseases and complications including those related with the age of 55 and older, stenocardia, fits, atherosclerosis, Charcot syndrome, diabetes, coronary disease, peripheral vessel disease, abnormal thrombocyte function, hemodialysis, hypercholesterolemia, hypertension, myocardial infarction, congestive cardiac failure, ischemia, nephropathy, high blood plasma homocysteine level, cardiac arrest or restenosis, diseases and complications associated with smoking, obesity and sedentary life-style causing a high risk of cardiovascular diseases in individuals, including apoplexy and paralysis.

EFFECT: preparing an effective drug for treating cardiovascular diseases.

28 cl, 3 dwg, 2 tbl, 5 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to new dihydropyran-2-one of formula

, where R1 is selected from hydrogen, ORa, OCORa, OCOORa, NRaRb, NRaCORb and NRaC(NRa)NRaRb; each R2 and R3 are optionally selected from hydrogen, substituted or unsubstituted C1-C12 alkyl, substituted or unsubstituted C2-C12 alkenyl and substituted or unsubstituted C2-C12 alkynyl; each R41, R42, R43, R44, R45, R46, R47 and R48 are optionally selected from hydrogen, substituted or unsubstituted C1-C12 alkyl, substituted or unsubstituted C2-C12 alkenyl and substituted or unsubstituted C2-C12 alkynyl; each R5, R6 and R7 are optionally selected from hydrogen, CORa, COORa, substituted or unsubstituted C1-C12 alkyl, substituted or unsubstituted C2-C12 alkenyl and substituted or unsubstituted C2-C12 alkynyl, or R5 and R48 together with coupled atom N and atom C whereto attached can form substituted or unsubstituted heterocyclic group; each Ra and Rb are optionally selected from hydrogen, substituted or unsubstituted C1-C12 alkyl, substituted or unsubstituted C2-C12 alkenyl, substituted or unsubstituted C2-C12 alkynyl, substituted or unsubstituted aryl and substituted or unsubstituted heterocyclic group; and each dashed line means an optional additional bond; to its pharmaceutically acceptable salts, tautomers or stereoisomers.

EFFECT: development of the method for preparing an anticancer composition, and the method of treating cancer.

46 cl, 10 tbl, 12 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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