The method of obtaining gamma-l-glutamylcysteine, the use of gamma-l-glutamylcysteine, the pharmaceutical composition

 

(57) Abstract:

Usage: in medicine. The essence of the invention: an improved method of obtaining gamma-L-glutamylcysteine (I) consisting in the acylation of histamine gamma panafcortelone ester L-tert.-butyloxycarbonyl--benzyl ester of L-glutamic acid in an organic solvent, followed by removal of protections in the acidic environment and hydrogenolysis, isoelectric precipitation and recrystallization. The use of compound (I) as a means to provide antioxidant, antiradical, operahouses, hypoglycemic, anti-asthma, hepatoprotective, antiviral, antibacterial, anticancer, antimetastatic, adaptogenic action, as well as the ability to modulate the arachidonic acid metabolism and other types of therapeutic action. Pharmaceutical composition having the above properties, including the active connection (I) in an effective amount and the target additives. 3 S. and 1 C.p. f-crystals, 14 PL.

The present invention relates to the field of Bioorganic chemistry and relates to improvements in a method of synthesis of L-glutamylcysteine (-L-Glu-HA), and goalside known pseudopeptide formula (I).

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-L-Glu-HA was first isolated in trace amounts from the nervous tissue of the rat and the clam [Koniski N., Kakimoto Y. /Formation of-glutamylhistamine from histamine in rat brain.// J. Neurochem. -1976. -vol. 27. -pp. 1461-1463; Tsuji M., Matsuoka Y., Nakajima T. /Studies of formation of-glutamines in rat brain and synthetic and catabolic enzymes.// J. Neurochemistry. -1977. -vol. 29. -pp. 633-638.]. Known chemical method get-L-Glu-HA, on the basis of Z-L-Glu(OEt) - OH [Koniski N., Kakimoto Y. / Formation of - glutamylhistamine from histamine in rat brain.// J. Neurochem. -1976. -vol. 27. -pp. 1461-1463]. The last action of hydrazine hydrate for 48 hours turn into the corresponding hydrazide Z-L-Glu(N2H3)OH with yields of 80%, and then in azide Z-L-Glu(N3)OH. The latter is introduced into reaction with histamine in aqueous-alkaline medium in accordance with the method of Kakimoto [Kakimoto Y. , Nakajima T. /Isolation of L-Glu-L-Glu acid and L-Glu-L-glutamine from bovine brain. // Biochemica et biophysica Acta. -1964. -vol. 93. -pp. 333-338] . The reaction product of Z-L-Glu(HA)OH hydronaut for removal of the Z-protection. Formed-L-Glu-HA purified column chromatography on an ion exchanger IR 120, followed by recrystallization from water, alcohol and acetone. Specific methods indicating the outputs of intermediate and final products and conditions of the separate stages of the synthesis of L-Glu-HA [Koniski N., Kakimoto Y. / Formation of - glutamylhistamine from histaimine in rat brain.// J. Neurochem. -1976, -vol. 27. -pp. 1461-1463] is not given.

The disadvantages of spooo free carboxyl group [Schroeder E. , Lyubts K. Peptides. M.-World. -1967. -S. 249] , the complexity of purification of the final product-L-Glu-HA [Koniski N., Kakimoto Y. / Formation of-glutamylhistamine from histamine in rat brain.// J. Neurochem. -1976. -vol. 27. -pp. 1461-1463], low yields and poor reproducibility azide method in the synthesis of related compounds: outputs-Glu--aminoisobutyric acid for the D-isomer is 17.7%, while for the L-isomer - 38,8. -L-Glu-HA synthesized by a known method [Koniski N. , Kakimoto y / Formation of-glutamylhistamine from histamine in rat brain. // J. Neurochem. -1976. -vol. 27. -pp.l461-1463], is characterized by only the elemental analysis and by acid hydrolysis, which is not enough to confirm the structure and identity of the substance, including the lack of it-isomer.

Known chemical method get-L-Glu-HA, on the basis of Boc-L-Glu(OH)-OButclose to the claimed and chosen as a prototype [Mc Caman M. W., Stetzler J., Clark C. / Synthesis-of-Glutamyilopamine and Other Peptidoamines in the Nervous System of Aplysia californica.// J. Neurochem. -1985. -vol. 45. -N-6. -pp. 1828-1835].

The disadvantage of this method is the length of the reaction off-OButbroadcasts from Boc-L-Glu(HA)-OButwhich, moreover, is accompanied by the formation of by-products. In addition, in the publication [Mc Caman M. W., Stetzler J. , Clark C. / Synthesis-of-Glutamyldopamine and Other Peptidoamines in the Nervous System of Aplysia californica.// J. Neurochem. -1985. -vol. 45. -N-6. -pp. 11), Boc-L-Glu(OPfp)-OBzl enter into interaction with the histamine in the environment of DMF.

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Intermediate Boc-L-Glu(HA)-OBzl purified by recrystallization and hydronaut in methanol over the catalyst is palladium on charcoal. Next otscheplaut Boc-protection triperoxonane acid. Isolation and purification of target-L-Glu-HA performed electrophoretic deposition under the action of an organic base in an environment of an organic solvent followed by recrystallization.

The advantages of this method are the high outputs, the practical absence of adverse reactions, exclusion from synthesis-low-tech stages of purification of the final and intermediate products column chromatography, conducting all stages of synthesis and purification in organic medium, since it is known that gamma-glutamina communication sensitive to the action of hydrolytic agents [Schroeder, E., lyubts K. Peptides. -M-World. -1967. -S. 249].

The original product Boc-L-Glu-OBzl commercially available and sold by the companies that produce reagents for peptide synthesis.

The method of synthesis described in the examples.

The individuality of the obtained compounds is checked by TLC on plates "Kieselgel" company "Merck" in the solvent system : chloroform-met is 4); on paper mill them. Street in the solvent system isopropanol-water-25% aqueous ammonia 6:3: 1(5).

Chromatogram showing chloro-toleenum solution, reagent Pauli and ninhydrin.

The melting point of a substance is determined on the device "Boetius (Germany).

1H-NMR spectra are removed on the instrument Bruker WM-250 (Germany) and Varian XL-400 (Japan) with TMS as internal standard.

Analytical reversed-phase HPLC performed under conditions (1): column IPU-270 C-18 (4.0 x 250 mm, 10 μm, elution of 0.1 M Na2HPO4, pH 2.3; (2) : the same column, elution of 0.1 M Na2HPO4, pH 2.7.

Example 1.

PENTAFLUOROPHENYL ESTER OF N-TERT.-BUTYLOXYCARBONYL- - BENZYL-L-GLUTAMIC ACID (II).

To a solution of 0.425 g (1.25 mmol) of Boc-L-Glu-OBzl in 6 ml of a mixture of dioxane and methylene chloride ( 1:1 ) was added 0.23 g (1.25 mmol) of pentafluorophenol in 1.5 ml of the same solvent mixture under stirring. Cooled to 0oC and add 0.258 g (1.25 mmol) of N,N'-dicyclohexylcarbodiimide. Stirred for 4 h, the precipitated DCU is separated. The solvent of the filtrate is removed in vacuo. To the oily residue was added 4 ml of hexane, pound. The precipitate was separated, dried in vacuum. Output 0.658 g-BENZYL-L - GLUTAMYLCYSTEINE (III).

To a solution of 0.250 g (0.48 mmol) of Boc-L - Glu(OPfp)-OBzl in 2.5 ml of DMF are added under stirring 0.055 g (0.48 mmol) of histamine, the pH of the reaction mixture is adjusted to 8 by addition of N-methylmorpholine. Stirred for 30 min at 20oC and leave for 48 hours at 0oC. the Dimethylformamide is removed in vacuo, the oily residue is triturated with 2 ml of dry ether. The precipitate was separated, washed with 2 ml of ether and recrystallized from acetone. Output 0.145 g (85.0%). Rf0.39 (2). So pl. 130-132oC. the IR spectrum (vaseline), cm-1: 1680 (amide I), 1530 (amide II).1H-NMR spectrum (CDCl3CD3OD), , M. D.: 1.4 (s, 9H, (CH3)3C-Boc), 2.1 (m, 2H, -CH2), 2.45 (t, 2H, -CH2), 2.85 (t, 2H, -CH2-HA), 3.45 (t, 2H, -CH2-HA), 4.2 (t, 1H, -CH), 5.2 (d, 2H, CH2-Bzl), 7.05 (s, 1H, CH-4-Im), 7.4 (s, 5H, C6H5-Bzl), 8.1 (s, 1H, CH-2-Im). Found %: C, 61.08; H, 6.85; N, 13.10. C22H30N4O5. Calculated %: C, 61.39; H, 6.97; N, 13.02.

Example 3.

-L - GLUTAMYLCYSTEINE (I).

To a solution of 0.050 g (0.11 mmol) of Boc-L-Glu(HA)-OBzl in 2 ml of anhydrous methanol was added 0.050 g of 10% palladium on charcoal, stirred for 2.5 hours, occasionally passing a current of hydrogen. The catalyst is filtered off, washed with 3 ml of methanol. The solvent of the filtrate is removed in vacuo. Get 0.040 g (100%) transparently ml of anhydrous ether. The precipitated oil was washed with ether when crushed and dried in vacuum over P2O5. The solid residue is dissolved in 0.4 ml of ethanol and add 0.036 ml Et3N to pH 7. Cheesy precipitate was separated, washed with ether and dried. Output 0.0235 g (89.0%). After recrystallization receive 17.1 mg (65.0%). Rf0.61 (4), Rf0.58 (5). HPLC under the conditions (4): single peak, time of release 3.50 min; (5): single peak, time of release 3.53 min1H-NMR-spectrum in D2O , M. D. : 1.8 (m, 2H, -CH2), 2.13 (t, 2H, -CH2/), 2.57 (t, 2H, -CH2-HA), 3.18 (t, 2H, -CH2-HA), 3.5 (t, 1H, -CH), 6.73 (s, 1H, CH-4-Im), 7.55 (s, 1H, CH-2-Im).

Biological activity.

Previously, based on the antioxidant properties of L-Glu-HA - known is the closest analogue of the claimed compounds was proposed to use it for the treatment of several diseases, in particular atherosclerosis, allergies [Seguin, M. S., Babizhayev, M. / Product de couplage de l'un acide amine, procede de preparation et applications therapeutigues et cosmetologigues.// Patent Fr. - 2701947. -1994. -C1 C 07 C 237/04. -A 61 K 31/195. -7/48; Babizhayev, M., Seguin M. C. / Pseudodipeptide product having on imidazole grouping and applications.// WO 95/12581. -C1 C 07 D 233/64. -C 12 P 17/10. -A 61 K 31/415. -7/42].

However, these indications have not been confirmed experimentally in experiments in vivo. It is shown that L-Glu-HA - inhibits the formation of superoxide anion only .With. / Pseudodipeptide product having on imidazole grouping and applications.// WO 95/12581. -C1 C 07 D 233/64. -C 12 P 17/10. -A 61 K 31/415. -7/42]. In addition, it is possible to predict greater instability-L-Glu-HA - to the action of enzymes in vivo than the corresponding analogue.

Based on these data, we have extended the study of the biological activity of the claimed compounds (IV). As a result, it was revealed previously unknown types of pharmacological activity and the mechanism of action on the basis of in vitro experiments. Thus, it was shown that the claimed compound has antioxidant and antiradical activities at low concentrations.

The combination of these properties determines the value of the proposed compound (IV) as pharmacological tools for the treatment of several diseases.

Examples of various kinds of biological activity shown by the claimed compound in vitro and in vivo is shown below.

Example 4.

EFFECT-L-Glu-HA HA the FORMATION of ACTIVE FORMS of OXYGEN IN MODEL SYSTEMS.

We also studied the effect of drug-L-Glu-HA change chemiluminescence (CHL), due to the formation of hydroxyl radical (.OH) and superoxide anion radical is) of different nature were generated in the following systems:

A. hydroxyl radical in the mixture FeSO4with H2ABOUT2(Fenton's reagent) [Halliwell Century / Superoxide-dependent formation of hydroxyl radicals in the presence of iron salts.// FEBS Lett. -1978. -vol. 96. -pp. 238-241].

The incubation medium consisted of 5 mm KH2PO4(pH of 7.4), 5 mm FeSO4; 2 mm lyuminola. H2ABOUT2at a final concentration of 5 mm was injected into the cell through dispender after it was recorded background glow of the mixture of reagents. Patent-pending compound, dissolved in water in the desired concentration was introduced into a cuvette in a volume of 5-10 μl. The total volume of the sample was 0.5 ml.

B. superoxide anion radical in the mixture of xanthine and xanthine oxidase [Afanasev I., T. Suslova, Cheremisina Z. et al. / Study of antioxidant properties of metal aspartates.// Analyst - 1995. -vol. 120. -pp. 859-862].

The incubation medium contained the following ingredients: 5 mm KH2PO4; of 0.2 IU/ml of xanthine oxidase. Xanthine concentration of 1 mm was injected into the sample through dispender. As a sensitizer glow in the system used lucigenin (0.2 mm). The test substance was administered similarly as in the study of hydroxyl radical.

In preliminary studies it was shown that the claimed compound did not affect the activity of xanthine oxidase.

is dispender) mixing. Indication signal chemiluminescence was carried out by integrating every 10 seconds - 5 minutes Duration of registration flash chemiluminescence after mixing of the ingredients was determined by the kinetics of a particular process. For each system were determined sutasoma chemiluminescence (mV) in control samples without drug (I) and in samples in the presence of appropriate concentrations of the drug (I+). To assess the degree of inhibition (activation) of CHL in the studied systems were (I+/I- (relative units).

The formation of ROS and the influence on this process for the proposed compounds were recorded on a device Luminometer-1251 (LKB, Sweden).

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

A. the Effect connections-L-Glu-HA on the formation of hydroxyl radicals in the Fenton's reagent.

With the introduction of H2O2in phosphate buffer containing ferrous sulfate, there was a flash of CHL, which was caused by formation in the reaction mixture predominantly hydroxyl radical. Flash was of limited duration, maximum intensity was reached at the 10th second and in the next 10-20 seque research actions-L - Glu-HA on the generation of hydroxyl radicals in the Fenton's reagent. The results show that the compound inhibits formation of hydroxyl radical in the studied system.

B. Effect-L-Glu-HA on the formation of superoxide anion radical in the system xanthine-xanthine oxidase.

Introduction xanthine in a medium containing xanthine oxidase, lucigenin, led to an outbreak of CHL, which reached a maximum within 3-5 minutes after which the system under investigation was started a very slow decrease in the intensity of the chemiluminescence. Adding-L-Glu-HA in a mixture of xanthine - xanthine oxidase has not fundamentally changed the shape of the curve PI response varied only the values of the maximum intensity of the chemiluminescence. As a result of this "stretching" of the kinetic curve was recorded sutasoma chemiluminescence for the first 3-5 minutes, which reflected the total number of light quanta produced in the system to achieve the maximum intensity of the chemiluminescence.

As can be seen from the presented data (table. 2), -L-Glu-HA has the ability to inhibit the formation of superoxide anion radical.

Example 5.

STUDY of ANTIOXIDANT ACTIVITY of L-Glu-HA EXPERIENCE IN VIVO.

The experiment was conducted on 47 outbred rats-samajpati) in animals caused by the introduction of carbon tetrachloride (CCl4) intragastrically in the form of a 50% solution in liquid paraffin in a volume of 0.25 ml per 100 g of body weight in 3 days [vengerovsky A. I., Chuchalin B. C., Pauwels O. C., Saratikov A. C. / Influence hepatic metabolism of lipids at CCl4- hepatitis.// Bull. the experts. Biol. -1987. - #4. -S. 430 to 432] . -L-Glu-HA was injected animals intragastrically at doses of 50 and 500 μg/kg to liver damage (3 and 4) for 3 days, and at the same time with CCl4(5 and 6 groups). Animals of the control group were injected CCl4as described above (group 2). Intact animals were treated orally with saline solution in an equivalent amount (group 1).

Samples of blood and liver were taken for analysis after 18 hours after the last injection of CCl4.

The content of the primary products of lipid peroxidation (LPO) - diene conjugates, diene ketones and tranov, was determined by the method [Valigorsky I. A., Nalimov A. G., Yarovinsky, B., Lifshitz, R. I. /Comparison of different approaches to the definition of products of lipid peroxidation in heptane - isopropanol extracts blood.// The matters. the honey. chemistry. -1989. -N 1. -S. 127-131]. The calculation of LPO products was performed by matching the values of the corresponding extinction to 1 ml of the investigated samples.

The experimental results were processed statistically using t-student criterion [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The results of table 3 indicate that the rat CCl4led to the accumulation of LPO products in the serum and liver. Animals in all groups, which were introduced-L-Glu-HA, there was a decrease in MDA content in serum and liver. In addition, the detected decrease in primary peroxidation products in the liver of animals treated with investigational compound.

Example 6.

EFFECT-L-Glu-HA ON Metamesh male line C57B1, were on a standard diet of the vivarium. The lungs of the animals were removed, frozen in liquid nitrogen, then homogenized with a glass homogenizer firm Wheaton (USA) at +4oC in 10 volumes of 0.05 M Tris-HCl buffer. Aliquots (0.5 ml) of the supernatant were incubated with 0.5 mccu [C14] -arachidonic acid ([C14]-AK, Amersham, England; specific activity 50-60 ICJ/mmol) at +37oC for 30 minutes Concentration in the incubation media-L-Glu-HA was 10-4M Extraction nematerializiranih [C14]-AK and products of its metabolism was carried out in 20 volumes of a mixture of chloroform and methanol (1:1), extraction efficiency not less than 90%, estimated using [C14]-PG F2. The solvents were removed on a rotary evaporator ("Buchi, Switzerland) under reduced pressure, the residue was dissolved in a mixture of chloroform: methanol (1:1). Separation and identification [C14] -AK and its metabolites was carried out by thin-layer chromatography on plates Kieselgel 60 (Merck, Germany), using organic phase solvent system (ethyl acetate, isooctane, acetic acid, water in a ratio of 110: 50:20:100), and labeled standards. Autoradiographically obtained on x-ray film X - Omat AR (Kodak, USA) and HS 11 ("ORWO", Germany) is conducted using radiometry fractions, received high performance liquid chromatography (HPLC-system firm "Gilson, France; column C8 BOND of "Du Pont" company USA) and elution of the spots on the TLC plates.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The results of the experiment are presented in table N-4. Noteworthy is the reduction in the number of PG f239%, 6-keto-PG f139% and 12-, 15-DECLINED by 28% in the presence of L-Glu-HA.

Thus, the introduction in the incubation medium connection L-Glu-HA leads to a change in the metabolism of arachidonic acid in the in vitro system.

Example 7.

Investigation of the EFFECT-L-Glu-HA ON the manifestations of EXPERIMENTAL BRONCHOCONSTRICTION.

Anti-asthma effect-L-Glu-HA was studied on the model of antigen-induced bronchoconstriction in actively sensitized Guinea pigs according to the method of Andersson [Andersson P. /Antigen induced bronchial anaphylaxis in actively sensitized guinea pig.// Allergy. -1980. -vol. 35. -pp. 65-71].

This model is the most adequate atopic bronchial asthma, as bronchospasm in Guinea-pig develops as a result of allergic reactions between antigen and homocytotoxin injection of 0.5 ml of suspension, containing 20 μg of ovalbumin (BOTH manufactured by Sigma /grade III/) and 100 mg Al(OH)3on the animal. The resolving dose of 150-200 mg/kg BOTH were injected intravenously (v. jugularis) in 0.1 ml saline on day 26 after sensitization. Induction of bronchospasm and measurement of parameters of external respiration was carried out according to the method described in [Yu-Hong L., Barnes, P., Rogers D. / Inhibition of neurogenic plasma exudation and bronchoconstriction by a K+chennel activator, BRL 38227, in guinea pig airways in vivo.// Europ. J. Pharmacol. -vol. 239. -pp. 257-259]. The animal was narcoticyou intraperitoneal introduction of etamine sodium (70 mg/kg), expose the trachea was performed tracheotomy and was inserted into the trachea cannula. The cannula was attached through a special tee to the apparatus (Ugo Basel, model 7025), who worked during the experiment in continuous mode: the volume of ventilated air 8 ml, respiratory rate of 70 per minute. Measurement of respiration parameters was carried out using the transducer (Ugo Basel, model 7020) connected to the cannula and recorder (Milligram) that record the amplitude of breathing. The amplitude reflected the degree of resistance of the smooth muscles of bronchial air flow. After the establishment of the Guinea pigs normal rhythm of breathing, the animal was injected in v. jugularis permits the dose Antiga is stvie their narrowing) and amplitude of respiration in 8-10 times compared with the original value. The dynamics of bronchospasm was observed within 30-60 minutes. When evaluating the effectiveness of the claimed compounds was determined by the change in the value of bronchospasm.

The test substance was dissolved in saline and was administered to three animals for 72, 48 and 18 hours before the induction of bronchospasm intragastrically at a dose of 50 mcg/kg as the comparison drug used Intal drug, have been widely used in the treatment of bronchial asthma. Intal animals were injected intragastrically at a dose of 5 mg/kg according to the same scheme as-L-Glu-HA. Control group animals received an equivalent amount of saline.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The data in table 5 show that the claimed compound significantly reduces the magnitude of bronchoconstriction by 40%, compared with the control values.

Thus, -L-Glu-HA showed activity towards the reduction of the magnitude of bronchoconstriction, comparable to the effect of the drug comparison - Intal. However, the current proposed dose of the substance was two orders of magnitude below stepnote-L-Glu-HA was studied on the model of experimental hyperlipidemia [Arichi N, Kumura H.About. / Effects of stibens compounds of roots of polygonum cuspidatium on the lipid metabolism.// Chem. Pharm. Bull. -1982. -vol. 30. -N 5. -pp.l766-1767] mongrel of male rats with initial weight 220-250 g, treated for 10 days intragastric against the standard diet in an oil suspension containing 10% cholesterol and 1% holeva acid (1 ml of suspension per 100 g of body weight). The investigated compound was administered to the animals orally at doses of 50 and 500 mcg/kg in the last four days of the experiment. As the comparison drug used nicotinic acid, which was introduced animals within 10 days on the background of the atherogenic burden in a dose of 10 mg/kg blood Samples were taken for analysis after 18 hours after the last injection of the drug, during which rats were deprived of food.

Was determined by the following parameters: total cholesterol (LDL - General), cholesterol high density lipoprotein (HDL-C), cholesterol of low-density lipoprotein and very low density lipoprotein (LDL-C and LDL-VLDL), triglycerides (TG). The content of cholesterol in blood serum was determined by the method Ilka [Biochemical studies in the clinic.- under. Ed. by A. A. Pokrovsky. -M-Medicine. -1969. -S. 300-302], HDL-C was evaluated in the supernatant after heparin-manganese precipitation of LDL+Leonia cholesterol-LDL.// Lab. case. -1979. -N 1. -S. 36-41]. Determination of LDL-C was performed by calculating according to the formula presented in the work of W. T. Friedewald et al [W. T. Friedewald, Levy K. J., R. Leus /Fat transhort lipoproteins in an integrated approach to mechanism and disoders.// New Eugl.J. Med. -1967. -vol. 276. -p. 32].

To assess the impact of the proposed connection on the ratio of atherogenic and antiatherogenic lipoproteins of blood plasma cholesterol was calculated index (Kxcthe formula proposed by Klimov A. N. with co-authors [Klimov A. N. , Nikulicheva N. G. Lipoproteins, dyslipoproteinemia and atherosclerosis. -M-Medicine. -1984. -165 C.].

The content of triglycerides in serum were determined by standard method [Radionova L. P. /Modification method determination of triglycerides in serum. / LBA. case. -1980. -N 5. -S. 297-299].

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The data given in tables 6, 7, suggests that the introduction of animal fat suspension was accompanied by a significant increase in the content of cholesterol in blood serum due to the atherogenic lipoprotein fractions (LDL and VLDL) on the background of reducing the number of atherogenic fraction - PL is 2">

Table 7 presents results of the effects of the studied compounds at a dose of 50 µg/kg lipid composition of the blood in animals fed the atherogenic burden. It is shown that the introduction of a-L-Glu-HA was changing all the investigated parameters almost to control values. In animals that were administered the compound at a dose of 500 mcg/kg, revealed similar changes.

Thus, changes in the lipid composition of serum introduction experimental animals studied compounds comparable to the lipid-lowering effect of the drug compared with nicotinic acid (table. 6). However, the effective dose of L-Glu-HA was two orders of magnitude lower than the comparison drug, and investigated the connection was administered within 4, not 10 days as nicotinic acid.

Example 9.

EXPERIMENTAL study of the HYPOGLYCEMIC ANTIDIABETIC ACTIVITY of L-Glu-HA.

Studies conducted on rats male Wistar weighing 250-300 g Experimental diabetes was induced by single intravenous streptozotocin (RSCN. "Synthesis" when IMG Academy of Sciences of Ukraine) at a dose of 42 mg/kg in rats, pre-starving within 24 hours from admission to food immediately after the injection. Rats were deprived of their 4 days in daily doses of 50 mg/kg and 500 mg/kg in aqueous solution at a ratio of 1 ml per 200 g of body weight. Control animals received a corresponding volume of water. Animals were deprived of food for 2 hours before blood sampling. The effect was estimated by the change of the glucose level in the blood after 2, 5 and 24 hours after the last injection of the drug. Blood was taken from tail vein in a volume of 0.1 ml of the glucose Content was determined by o-toluidine method.

The glucose content was calculated in mg% on a calibration curve using standard solutions of glucose. Next, we determined the rate of change of glucose in relation to the original number for each animal, expressing it in percentage of the initial content. The final calculation was performed for each group of animals.

The results of the experiments are processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The data in table 8 show that 4-day intragastric administration of L-Glu-HA in daily doses of 50 and 500 mg/kg resulted in a significant reduction in blood glucose 2 hours after the injection the animals. Hypoglycemic effect remained after 5 hours after administration, however, was less pronounced and not statistically significant.

So obrw a dose of 50 mg/kg, and 500 ág/kg

Example 10.

EFFECT-L-Glu-HA FOR the MAINTENANCE of BLOOD GLUCOSE IN the INTACT ANIMALS.

The experiment carried out on outbred rats male with the original weight 200-220, Animals of the control group (group 1) for 3 days were treated orally with saline, the animals of the experimental groups (2 and 3 groups) - solution-L-Glu-HA at doses of 50 and 500 µg/kg, respectively. On the day of blood sampling, animals were deprived of food. Two hours before the last injection of the drug in animals took the blood from the tail vein. Blood samples were taken after 2 and 5 hours after administration of the compound, then the animals were placed on a diet of the vivarium and the last blood sampling was performed 24 hours after the last injection of the drug. The glucose content in the samples was determined as described in example 7.

The results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The data presented in table 9, suggest that food deprivation resulted in a slight, but significant decrease of glucose in animals of all groups, followed by reduction to the number close to the original. Not willerson, introduction-L-Glu-HA did not affect the glucose content in the blood of intact animals.

Example 11.

ANTIVIRAL ACTIVITY of L-Glu-HA.

A. Protective effect of L-Glu-HA when the infection caused by the virus encephalomyocarditis.

Investigations were carried out on outbred mice of both sexes with initial mass 10-11, In the control and experimental groups were used for 30 animals. The investigated compound and a drug comparison of Radostin - interferon inducer, was injected once intraperitoneally 24 hours after infection of mice with virus encephalomyocarditis in doses of 30 mg/kg and 5 mg/kg, respectively. Virus encephalomyocarditis was administered at a dose of 100 LD50. Antiviral activity was determined from the change in life expectancy (ALE) mice and the degree of protection against lethal viral infection.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The results are presented in table 10. It is shown that L-Glu-HA had a pronounced antiviral activity at a dose of two orders of magnitude lower than the dose of the comparator drug.

B. Effect-L-Gl the breed mice of both sexes weighing 8-10, The non-human influenza virus, type A, strain Aichi (allatoona liquid), was administered to mice intranasally at a dose of 100 LD50, -L-Glu-HA was administered to animals orally at doses of 50 and 500 mcg/kg over 3 days before infection and 10 days after infection. As Comparators used Arbidol (specific drug) at a dose of 100 mg/kg, was administered for 24 and 2 hours before and 3 days after infection, and Timogen (non-drug) at a dose of 10 mg/kg, administered to animals for the same pattern as the test substance. In each group used for 30 animals. Antiviral activity was determined from the change in mean life span of mice and the degree of protection against lethal doses of virus infection.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

The results of the study are presented in table 11. It is revealed that the increase in mean life span, survival and the degree of protection is most pronounced with the introduction of the compounds at a dose of 500 mcg/kg Application-L-Glu-HA in a dose of 50 mg/kg and Arbidol was accompanied by change of the investigated indicators equally. Action Thymogen as antiviral drug was virusnom action of the claimed compounds in experimental viral infections in mice caused by the virus of human influenza type A. Effect-L-Glu-HA was comparable and somewhat more pronounced than Arbidol. However, the effectiveness of the claimed compounds marked with doses of 3-4 orders of magnitude lower compared to the reference preparation Arbidol.

C. Effect-L-Glu-HA on the reproduction of human immunodeficiency virus in acute infection lymphoblastoid cells.

The study was performed on the culture of lymphoblastoid cells MT-4. In the experiment used the human immunodeficiency virus type 1, HIV-1 isolate/IV from the collection of the Institute of Virology. D. I. Ivanovsky. -L-Glu-HA was tested in concentrations: of 1.0, 0.1 and 0.01 μg/ml Drug comparison - azidothymidine, at a dose of 0.05 μm/ml of the Substance brought into the culture of cells MT-4 for 1 hour before making the virus at a dose of 1000 TCID50.

The main parameter of the effectiveness of the compounds were cell viability.

In preliminary experiments it was shown that L-Glu-HA low toxicity to lymphoblastoid cells at concentrations of 1.0 μg/ml or less.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-N which was amadala pronounced protective effect from cytodestructive steps of HIV-1 in concentrations of 1.0 and 0.1 μg/ml, comparable to the effect of the drug comparison - azidothymidine.

Example 12.

EFFECT-L-Glu-HA ON the RESISTANCE of ANIMALS TO MICROBIAL INFECTIONS.

The experiment used a nonlinear white mice weighing 18-22 g-L-Glu-HA product comparison - Nukleinat sodium was administered orally for 3 days before and after infection (figure -3, -2, -1, 0, 1, 2, 3, where 0 is the day of infection). For infection of animals used daily culture of Salmonella sp., grown on agar of Hottinger. For preparation of cell suspension was used saline. Conditions of infection was tested in preliminary experiments. Cell suspension Salmonella sp. was administered subcutaneously at a dose of 5108cells/mouse in 0.5 ml-L-Glu-HA was administered orally at doses of 150 and 500 mg/kg, and the product comparison - Nukleinat sodium at a dose of 50 mg/kg in 0.5 ml saline.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

Introduction the compounds at a dose of 150 and 500 mg/kg significantly increased the average life expectancy infected with Salmonella sp. mice (table. 13), providing a protective effect in hard esteem Nucleinate sodium, a known stimulator of immune responses, including phagocytosis [Lazarev D. L. , Alekhin E. K. immune Stimulants. -1985. -286]. Survival of animals on the second day after infection in the group of mice treated with the test substance at a dose of 150 μg/kg, 70%, and at a dose of 500 mcg/kg is 80%, whereas in the control group only 50%.

Thus, -L-Glu-HA has a strong protective effect against microbial infection.

Example 13.

The STUDY of the effect-L-Glu-HA ON TUMOR GROWTH.

Postmodifiers effect of the drug was studied on the model of adenocarcinoma of the submandibular salivary gland CT-6. In the experiment mouse-female line DBA/2, obtained from the kennel "Pole". Strains tumor CT-6 transplanted suspension tumor tissue in medium 199 (1:10) subcutaneously. -L-Glu-HA in a dose of 50 mg/kg was administered parenterally, and the product comparison - indomethacin at a dose of 1500 mg/kg oral. The first injection of drugs was 24 hours after inoculation of the tumors. Introduction compounds to animals with tumor CT-6 -28 days.

The experimental results were processed statistically [Gubler E. C. Computational methods of analysis and recognition of pathological processes. -M-Science. -1978. -365].

Pokazal significantly lower compared to control animals.

Thus, -L-Glu-HA significantly reduces the growth rate of the investigated tumors.

The experimental results show high biological activity of compounds-L-Glu-HA.

As can be seen from the results (example 4), the claimed compound significantly inhibits the formation of hydroxyl radical and superoxide anion radical (O2-in vitro systems. Especially valuable is the fact that the rate of formation of hydroxyl radical (.OH), as in a living organism there is no enzymes, able to beat them. OK there are mechanisms for the effective removal of hydrogen peroxide and superoxide anion radical, which makes education.OH impossible [Hochachka P., Somero j. Biochemical adaptation. -M-World. 1988 at M.V.Lomonosov. -568 C.]. However, in stressful situations, the formation of hydroxyl radical, which, in turn, increases lipid peroxidation (LPO) and can cause damage to DNA.

The common link in the pathogenesis of a large range of pathological conditions (stress, radiation injury, allergic disease, atherosclerosis and related diseases, diabetes and others) is the activation of the FLOOR. This Pia membrane-bound enzymes. Normalization FLOOR can be an important option in the treatment of these diseases [Racer N. P., Manico centuries, Kirilovich L. M. Clinical pharmacology of hepatoprotectors. -Ternopil. -Zbruch. -1995. -272 C.].

It is known that the introduction of carbon tetrachloride (CCl4) leads to characteristic changes in the liver with signs patrilineage cirrhosis, increased lipid peroxidation and is a recognized model of liver injury accompanied by activation of the FLOOR [Kapil A., Koul LB., Suri O. R./ Antihepatotoxic Effects of Chlorogenic Acid from Anthocephalus - Cadamba.// Phytother Res. -1995. -vol.9. -Iss 3. -pp. 189-193]. Action CCl4is primarily due to damage to the endoplasmic reticulum, lysosomes and other membrane structures of cells due to activation of GENDER-induced formation of free radical metabolites CCl4[Vengerovsky A. I., Chuchalin C. O. , Pauwels O. C., Saratikov A. O. / Influence hepatic metabolism of lipids at CCl4- hepatitis.// Bull. the experts. Biol." 1987. - #4. -S. 430 to 432; Debt A. C., Dushkin M. I., Morozov, A. C., Morozov, I. Y. /change the content of lipids in the liver, blood plasma, aorta and activity cholesterolaemia rat liver when exposed to carbon tetrachloride.// The matters. the honey.chemistry. -1986. -N 1. -S. 55-5 8] . When porazeni which contributes to the accumulation of reactive oxygen species (including hydroxyl radicals) and the manifestation of their toxic effects on membrane structure of hepatocytes [Matyushin B. N., Loginov A. S., Pasha, S., Raslova E. M. /Enzymatic recycling of reactive oxygen species and lipid composition of the liver, with its toxic liver./ VI Symposium on the biochemistry of lipids. Proc. Dokl. -M. -1995. -S. 64].

Antioxidant and hepatoprotective effect-L-Glu-HA experience in vivo was studied on the model of liver injury with the introduction of animal CCl4(example 5). It is revealed that with the introduction of animals of the claimed compounds significantly reduced the content of malondialdehyde (MDA) in the serum in all groups of animals, and in the liver of rats treated with L-Glu-HA before the introduction of the CCl4. There is also a decrease in the liver of diene conjugates and diene ketones and trienol under the influence of the claimed matter.

The obtained results allow to recommend the use of-L-Glu-HA when the effects on the body such factors as stress and radiation, to prevent side effects during radiotherapy for activation of repair processes; in the treatment of liver diseases such as cirrhosis and hepatitis of different etiology; conditions associated with intoxication by substances which activate pareci is izvodstvo, associated with obtaining chemicals and contact with toxic substances.

It is known that the aging process is also associated with oxidative stress, accompanied by an increase in the FLOOR and, as a consequence, the formation of MDA, which, interacting with lysine, forms lipofuscinosis pigment that accelerates lipofuscinosis and causes irreversible morphological changes in the organs. The decrease in MDA content (example 5) reduces the probability of formation of this compound, which may lead to a decrease in the probability of developing age-related diseases. Oxidative stress in the nervous tissue (increase FLOOR) is one of the pathogenesis of several diseases of the nervous system, in particular Parkinson's disease. Complex geriatric diseases includes pathological conditions such as senile psychosis, cataracts, senile skin changes. Based on these data it is possible to offer the use of the proposed drug for the treatment of these diseases, and Parkinson's disease.

It is known that arachidonic acid (AA) is the precursor of many biologically active compounds, such as prostaglandins (PG) and lacunae metabolism AK (example 6). Thus there is a change in the ratio of PGE2and PG f2having the opposite effect on smooth muscles of the bronchi and blood vessels, which may serve as one explanation for the reduction in severity of experimental bronchoconstriction (example 7). In addition, the reduction of the synthesis of 12-NET is a factor that reduces the development of carcinogenesis [Honn, K. V., D. G. Tang, X. Gao , I. A. Butovich, B. Liu, J. Timar, W. Hagmann/12-lipoxygenases and 12(S)- HETE: role in cancer metastasis.// Cancer and Metastasis Reviews. -1994. -vol. 13. -Iss 3-4. -pp. 365-96. -Ref: 195].

As noted above, -L-Glu-HA has a strong antioxidant effect (examples 4, 5). The reduction in the severity of bronchospasm under the influence of the claimed compounds (example 7) pathogenetically may also be due to its antioxidant effect, as it is known that in patients with bronchial asthma in contact with the allergen, the accumulation of free radicals and activated lipid peroxidation (LPO). In remission showed a significant reduction in lipid peroxidation in these patients [Fedoseeva, B. Mechanisms of obstruction of the lungs. -Saint-Petersburg. -Medical news Agency. -1995. -334 C.-S. 129-132].

Thus, the results of the research allow recom>Lipid metabolism characterized by elevated levels of triglycerides, total cholesterol, cholesterol in the lipoproteins of low and very low density (LDL and VLDL) and decreased content of cholesterol in the lipoproteins of high density, leads to the development of such widespread diseases as atherosclerosis and obesity, and as a consequence of coronary heart disease, myocardial infarction, and is a risk factor for the manifestation of diabetes. It should also be noted that by reason of infringement of cholesterol metabolism may be the change of the structure of LDL and VLDL, as well as the activity of cell receptors. Found that patients with atherosclerosis and diabetes LDL cholesterol increased triglycerides and VLDL - cholesterol. On the other hand, it is shown that in individuals with a high content of glucose in the blood is the process of glycosylation of LDL, and with the increase of LPO products, for example, MDA formed peroxide-modified lipoproteins, primarily LDL. Last, due to its cytotoxicity can damage endothelial cover vessels. Was a positive correlation between lipid peroxidation in LPN the e high atherogenicity [Klimov A. N., Nikulicheva N. , Lipids, lipoproteins, and atherosclerosis. -Peter. -Saint-Petersburg. -1995. -302 C.].

It is known that changes in the content and ratio of lipids in plasma reflects changes in membrane structures parenchymatous organs. The composition of cell membranes, for example, microsomal, directly depends on the composition of the diet of experimental animals [Reichen J., Buters, J. T. M., Sojcic z, Roos F. J./ Abnormal Lipid-Composition of Microsomes from Cirrhotic Rat-Liver - Does It Contribute to Decreased Microsomal Function.// Experientia. -1992. -Vol. 48. -Iss 5. -pp. 482-486]. Introduction animal cholesterol causes its accumulation in cell membranes, reducing its fluidity, which in turn leads to changes in the functional state of enzymes [Wade A., Harred W. / Effect of dietary lipid on the drug-metabolising.// Feder. Proct. -1976. -vol. 55. -pp. 2475-2479; Buters, J. T. M., Zysset T., Reichen J./ Metabolism of Antipyrin in-Vivo in 2 Rat Models of Liver-Cirrhosis - Its Relationship to Intrinsic Clearance in Vitro and Microsomal Membrane Lipid Composition.// Biochem. Pharmacol. in 1993. -vol. 46. -Iss 6. -pp. 983-991].

Modeling of disorders of lipid metabolism, usually associated with the addition to the diet of animals of various fat compositions or the increased amount of cholesterol. Used in our research model of hypercholesterolemia (example 6) causes significant changes in the number and ratio of lipids in the blood: increased three the C-VLDL - "atherogenic" lipoproteins), cholesterol index (KXC) and the reduction of cholesterol high density lipoprotein (HDL-C is "antiatherogenic" lipoproteins). These changes in lipid composition of the blood are similar to the changes in patients with atherosclerosis, obesity and diabetes [Klimov A. N., Nikulicheva N. G. Lipids, lipoproteins, and atherosclerosis. -Peter. -Saint-Petersburg. -1995. -302 S.; Taton J. N. Obesity pathophysiology, diagnosis, treatment. -Warsaw. -Polish Medical publishers. -1981. -364 S.; Balabolkin M. I. diabetes mellitus. -M-Medicine. -1994. -384 C.].

When studying the efficacy of L-Glu-HA on the lipid metabolism revealed (example 8) significant reduction in the content of triglycerides, total cholesterol, LDL-C, LDL-VLDL and KXCand increasing the amount of HDL-C. It should be specially emphasized the importance of the fact that under the influence-L-Glu-HA there was a significant increase in HDL, as it is known that this lipoprotein fraction has a protective effect in the development of atherosclerosis and coronary heart disease. The protective effect of HDL is associated with the fact that they are involved in the transport of cholesterol from tissues to the liver, reducing its accumulation in the vascular wall [Ohkuwa T. , Sato Y., Naoi M /Hydroxyl Radical Formation in Diabetic Rats Induced by Streptozotocin/Nikulicheva N. G. Lipids, lipoproteins, and atherosclerosis. -Peter. -Saint-Petersburg. -1995. -302]. In addition, HDL prevents the formation of blood clots on the surface of atherosclerotic plaques. HDL can also block the LDL receptor, and then reduces the accumulation of cholesterol in cells. The experiment shows that even with an increased content of cholesterol in blood serum in the presence of high amounts of HDL prevents the formation of atherosclerotic plaques in blood vessels [Ohkuwa T., Sato Y., Naoi M /Hydroxyl Radical Formation in Diabetic Rats Induced by Streptozotocin// Life Sci. -1995. -Vol 56. -Iss 21. -pp. 1789-1798.].

We would like to highlight the decrease in the content of triglycerides in the serum with the introduction of L-Glu-HA, as it is known that TG may be an independent risk factor for coronary heart disease, without any toxic effect on the vascular endothelium, leading to increased platelet aggregation and reduced fibrinolytic activity of blood [Ohkuwa T., Sato Y., Naoi M /Hydroxyl Radical Formation in Diabetic Rats Induced by Streptozotocin// Life Sci. -1995. -Vol 56. -Iss 21. -pp. 1789-1798].

Operahouse effect-L-Glu-HA, apparently, can be associated with its antioxidant properties (examples 2, 3) as lipid metabolism, as a rule, is accompanied by an increase in the FLOOR.

Thus, -L-Glu-HA may be the Arda), obesity and diabetes for the prevention of blood clots.

In further studies it was shown that L-Glu-HA significantly reduces the amount of glucose in the blood in animals with experimental streptozotocin diabetes (example 9) without changing this indicator in healthy animals (example 10). Apparently, the decrease in the amount of glucose in the blood of animals with experimental streptozotocin diabetes is associated with normalization of lipid metabolism and reduction of lipid peroxidation in these animals, as well as the ability-L-Glu-HA to reduce the formation of hydroxyl radical, which plays an important role in the formation of the pathological process /39/. The effect of the drug is probably breaking observed in diabetes mellitus, when changing the metabolism of carbohydrates leads to a disturbance of lipid metabolism and Vice versa, since there is a close relationship of these two sides of metabolism in the body [Sokolov, I. E. diabetes mellitus and atherosclerosis. -M-Science. -1996. -405 C.].

Thus, these results confirm that L-Glu-HA stabilizes the homeostasis of the organism, which gives reason to recommend it for the treatment of diabetes.

Results viral infections (virus encephalomyocarditis and influenza virus), and also exerts a protective effect in cell cultures from cytodestructive steps of human immunodeficiency virus - HIV-1/HIV. Antivirus action-L-Glu-HA may be due to its antioxidant properties, as previously identified that substances with this action, have antiviral properties [Orlova T. G., F. Voronin Century , Nikiforov, A. , Ershov centuries, Burlakova E. B. /Antiviral activity of drugs with antioxidant properties.// III Ross. NAT. Congress. -Man and medicine. -M. -1996. -C. 41].

In addition, the data given in example 12 show that-L-Glu-HA significantly increases the life expectancy of animals infected with Salmonella sp. and treated with the investigational compound.

Thus, the proposed drug can be recommended for the treatment of bacterial and viral infections, including HIV.

It was revealed that the introduction of a-L-Glu-HA animals with transplantable tumor leads to a significant reduction in the growth rate of the studied tumors (example 13). It should be noted that the patented connection affects tumor growth similar to the drug compared to Indomethacin, which is used in practical medicine when l is flammatory treatment vay prolong survival in undernourished panients with metastatic solid tumors.// Cancer Res. -vol. 54. -N 21. -pp. 5602-5606], but the dose of patented compounds was in the order below.

It is known that the inhibition of the synthesis of 12-NET leads to a decrease in the interaction of tumor cells with endothelial cells of blood vessels and reduce the process of hematogenous metastasis [Honn, K. V., Tang, D. G., Grossi I., Duniec Z. M., J. Timar, Renaud C., Leithauser m, Blair I., Johnson C. R., C. A. Diglio /Tumor cell-derived 12(S)- hydroxyeicosatetraenoic acid dosage microvascular endothelial cell retraction.// Cancer Research. -1994. -vol. 54. -pp. 565-74; Honn, K. V., I. M. Grossi, C. A. Diglio, Wojtukiewicz, M., Taylor, J. D./Enhanced tumor cell adhesion to the subendothelial matrix resulting from 12(S)-HETE-induced endothelial cell retraction.// Faseb J. -1989. -vol. 3. -pp. 2285-2293. ] so the claimed compound may have antimetastatic properties.

In some works it is shown that the clinical use of drugs with antioxidant activity leads to improved functioning or normalization of the immune and metabolic systems of homeostasis in cancer [Kolomiets L. A., Suslov Ie /antioxidant therapy as a correction immuno-metabolic disorders in patients with precancerous diseases of the stomach. // II Russian national Congress "Man and medicine". J. struct. Dokl. -1995. -S. 199.; Laktionov K. L., Science Century. N. /Value of antioxidant therapy in oncogynecology is my connection with cancer to reduce the growth rate of the tumor and the extent of metastasis.

A wide range of actions offered drugs may be partly explained by the fact that-L-Glu-HA acts on certain systems, stabilizing the homeostasis of the organism. It is known that in vivo changes in the activity of one system leads to other changes and these changes take the cascading nature. With the introduction of the claimed compounds in the body comes normalization of a number of functions that allows you to take it to the adaptogens.

In conclusion, it should be noted that-L-Glu-HA exhibits a biological activity in doses of two to three orders of magnitude lower than Comparators with almost the same efficiency. Along with this, in the study of toxicity was not found sredneseriynoe dose, as even the introduction of animal-L-Glu-HA in a dose of 5000 mg/kg oral and 10000 µg/kg parenteral does not lead to death of the experimental animals.

-L-Glu-HA can be administered by intramuscular, intravenous, intranasal, oral, sublingual, intrarectal and applied in the form of inhalation. Accordingly, it can be used in various pharmaceutical forms such as injectable solutions, tablets, suppositories, ointments. Using the proposed connection is body mass). As diluent compounds may be used in 0.9% sodium chloride solution, distilled water, a solution of novocaine injection, ringer's solution, glucose solution for injection and others. -L-Glu-HA in the form of tablets and suppositories contain the active ingredient in the amount of 3.0-6.0 mg per tablet or a suppository. For tablets and candles as a pharmaceutical excipient use any pharmaceutically suitable basis. Patented connection in any form you can apply for 5-10 days 1-2 times a day. -L-Glu-HA has no side effects and no contraindications for use. Prepare dosage forms of the inventive compounds commonly known method.

Based on the results of the research suggested the use of-L-Glu-HA for the treatment and prevention of the following conditions and diseases:

1. stress;

2. bronchial asthma;

3. atherosclerosis, coronary heart disease, obesity;

4. diabetes mellitus type 1 and 2;

5. alcoholism, alcohol intoxication;

6. prevention of blood clots;

7. hepatitis, cirrhosis of the liver;

8. toxic liver damage;

9. diseases of the nervous system, Parkinson's disease;

10. gerontophilia radiotherapy;

12. viral and bacterial infections, in particular HIV;

13. cancer.

The list of abbreviations.

AK - arachidonic acid

ROS - reactive oxygen species

HPLC - high performance liquid chromatography

LD - lethal dose

LO lipoxygenase way

LPWN - high-density lipoprotein

LDL - low density lipoprotein

VLDL - very low density lipoprotein

LT - leukotrienes

MDA - malonic dialdehyde

OVA - chicken ovalbumin

PG - prostaglandin

FLOOR - lipid peroxidation

ALE - life expectancy

TBA - thiobarbituric acid

TG - triglycerides

THU - trichloroacetic acid

TZID tissue cytopathic dose

F - phospholipids

FR - saline

CHL - chemiluminescence

LDL - cholesterol

But- tert-butyl

Bzl is benzyl

Boc - tert-butyloxycarbonyl

DCC is N,N'-dicyclohexylcarbodiimide

DCU - N,N'-dicyclohexylmethane

DMF - N,N'-dimethylformamide

Et3N - triethylamine

EtOH - ethanol

Glu - glutamic acid

HA - histamine

5-HETE and 5(S)-hydroxy-who

OPfp - pentafluorophenyl

TFA - triperoxonane acid

Z - benzyloxycarbonyl

1. The method of obtaining gamma-L-glutamylcysteine by acylation of histamine substituted derivatives of L-glutamic acid and subsequent removal of protections, characterized in that as Alliluyeva agent use gamma-pentafluorophenyl ester of N-tert.butyloxycarbonyl--benzyl ester of L-glutamic acid and the process is conducted in an organic solvent, followed by removal of the benzyl protecting the hydrogenolysis over a palladium catalyst in an organic solvent and tert.-butyloxycarbonyl protection in acidic medium, with subsequent electrophoretic deposition of organic solvent and recrystallization.

2. The method according to p. 1, characterized in that the acylation reaction is carried out in dimethylformamide, cleavage of the BOC-protectors triperoxonane acid, and the solvent during isoelectric precipitation of applied ethanol.

3. The use of gamma-L-glutamylcysteine as a means to provide antioxidant, antiradical, operahouses, hypoglycemic, anti-asthma, anti-viral, antibacterial, protivoanemi acid, to prevent signs of diabetes, obesity, coronary artery disease, stress, hepatitis, cirrhosis, toxic liver damage, alcoholism, radiation injuries, geriatric changes.

4. Pharmaceutical composition having antioxidant, antiradical, operahouses, hypoglycemic, anti-asthma, anti-viral, antibacterial, anticancer, antimetastatic, adaptogenic action, as well as the ability to modulate the arachidonic acid metabolism, prevent signs of diabetes, obesity, coronary artery disease, stress, hepatitis, cirrhosis, toxic liver damage, alcoholism, radiation injuries, gerontological changes, containing as the active ingredient gamma-L-glutamylcysteine or its pharmaceutically acceptable salt in an effective amount and the target additives.

 

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