Copolymers of hetero-chain aliphatic poly-n-oxides, based on them vaccinating and medicinal preparations

FIELD: medicine.

SUBSTANCE: copolymers of hetero-chain aliphatic poly-N-oxides of general formula (I) , where R=N, CH; x=2-4; y=0, 2; n=10-1000; q=(0.1-0.9)n; z=(0.1-0.9)n. Copolymers possess anti-oxidant action, therapeutic action as detoxicant and immunomodelling agent. Copolymers of formula (I) can be used as immunomodulating carrier for obtaining vaccinating medication and as carrier of medications for obtaining medications.

EFFECT: copolymers of hetero-chain aliphatic poly-N-oxides represent novel class of compounds possessing wide spectrum of pharmacological and vaccinating action, aimed at increase of safety in application, increase of technological and economical effectiveness and ecological safety of production of medications.

20 cl, 2 dwg, 13 tbl, 22 ex

 

The invention relates to the field of synthesis of new high-molecular chemical compounds possessing biological activity, and can be used in the production of highly effective pharmaceutical drugs and vaccines.

Known synthetic polymers having physiological activity, for example, Homo - and copolymers of acrylic and methacrylic acid, copolymers of N-vinylpyrrolidone with acrylic and crotonic acids, acrolein, vinylamine, maleic anhydride and other vinyl monomers Polymers in medicine, translation from English. Ed. Board, N.A., M., 1969, p.38-76). Also known some water-soluble derivative chain polymers (plate N.A., Vasiliev A., "Physiologically active polymers", M., Chemistry, 1986, p.12-204; "Polymers for medical purposes", edited by Senoo Meno, translated from jap. M., 1981, 248 pages)containing nitrogen in the side chain of the macromolecule, such as polyvinylpyridine or polyvinylcarbazole. All the above compounds are toxic and are not widely used as pharmacological agents. In addition, chain polymers do not dissolve in the body for low-molecular compounds accumulate and cause unwanted side effects.

Most similar compounds on the biological effects and chemical nature of the described image is the shadow are derivatives of poly 1,4-ethyleneimine (RF patent No. 2073031, MCI C08G 73/02, publ. 1997). The known compounds are characterized by a wide spectrum of pharmacological actions and successfully find use as carriers of vaccine antigens or drugs, as well as in the treatment of diseases associated with immunodeficiency States. At the same time, the presence of ionic groups in these compounds reduce the rate of degradation and excretion from the body, which limits their use for intravenous administration as a detoxifier in the rapid excretion of toxic compounds. In addition, the technology of production of known compounds involves a multistep process associated with high consumption of expensive chemical reagents, requiring significant energy capacity and labor costs.

The technical result of the present invention is to create a new class of compounds possessing a wide spectrum of pharmacological and wakciniruemogo action with antioxidant and detoxication properties and are safe to use, as well as to improve manufacturability, cost-effectiveness and environmental safety of production of drugs.

This technical result is achieved by the compounds representing the copolymers of heterochain Ali is oticheskih poly-N-oxides of the General formula (I):

where R=N, CH;

x=2-4; y=0, 2; n=10-1000; q=(0,1-0,9)n; z=(0,1-0,9)n,

possessing pharmacological activity.

is a copolymer of triethylenediamine and N-oxide of triethylenediamine.

The copolymer of formula 1 has an antioxidant effect.

The copolymer of formula 1 has a therapeutic effect and can be an antidote.

The copolymer of formula 1 has a therapeutic effect and may be an immunomodulatory agent.

The copolymer of formula 1 can be immunoadjuvants.

The copolymer of formula 1 may be immunomodulatory media antigen or carrier medicinal substance.

This technical result is also achieved by the fact that vaccinifolia means comprising an antigen and an immunogenic carrier, contains as an immunomodulating media copolymer of formula 1.

This vaccinifolia the tool may be a conjugate of the antigen with the copolymer under item 1 in the presence in the structure of the antigen reactive functional groups.

Vaccinifolia the tool may be a compound obtained by the complexation of the antigen with a copolymer of formula 1.

Vaccinifolia tool can be a composition, obtained in the result is the mixing of the antigen with a copolymer of formula 1.

This technical result is achieved by the fact that the vaccine against hepatitis a and hepatitis "B" is characterized by the fact that it contains a vaccine formulation containing both AG CAA and HBsAg or vaccine preparations against hepatitis a and hepatitis b and copolymer hetero-chain aliphatic poly-N-oxides according to claim 1.

While a vaccine against hepatitis a and hepatitis "B" may contain as AG HAV antigens derived from strain FOREHEAD-86 hepatitis a virus in the culture of transplantable cells 4647, and the content of components in the dose of

AH HAV40-60 ELISA units
HbsAgthe 2.5-20 mcg
SP-MO0,1-10 mg

This technical result is also achieved by the fact that vaccine composition comprising the vaccine formulation and immunoadjuvant, contains as immunoadjuvant copolymer according to claim 1.

This technical result is also achieved by the fact that the drug, including drug substance and the medium contains as a carrier of the copolymer of formula 1.

The drug may be a conjugate of medicinal substance with a copolymer of formula 1 when the availability is in the structure of the drug substance reactive functional groups.

The drug may be a compound obtained by the complexation of drug substances with a copolymer of formula 1.

The drug may be a pharmaceutical composition obtained by mixing a medicinal substance with a copolymer of formula 1.

Copolymers hetero-chain aliphatic poly-N-oxides according to formula 1 (SP-NO) have pharmacological activity and, above all, show a pronounced antioxidant and detoxification properties, immunomoduliruuchee, antiviral and antibacterial activity, biogenic, as are analogues of natural products of oxidative metabolism natural hetero-chain polyamines and applicable to create safe for a living body of drugs.

SP-NO will biodestruction in the body on oligomeric and low molecular weight compounds and completely excreted. Thanks to its physiological activity-based compounds in accordance with the invention can be created by high ballastless pharmacological systems. Available reactive tertiary nitrogen atoms in the described connection opens wide possibilities of modification for the introduction of new reactionactos the groups in the side chain SP-NO with their further use as a carrier for conjugated vaccines, enzymes of various drugs. Obtaining conjugated compounds based on SP-NO possible only in the presence of reactive groups in vaccine antigens, enzymes, and drugs.

The described copolymers are characterized by a unique adsorption ability, which is caused by a combination of physico-chemical properties of these compounds, namely:

- a wide range of molecular weights, including high molecular weight;

- the presence of many weakly charged N-oxide groups in the main chain of the macromolecule and, as a consequence, the ability of the education of the equilibrium electrostatic complexes with other molecules;

- high polarity (dipole moment of N-O bonds is about 5 D, which is almost an order of magnitude larger than the dipole moment of the other links), which provides the formation of a stable electrostatic macromolecular complexes;

the formation of chelate complexes with metals and, as a consequence, the effective protection of cell membranes.

SP-NO have in the structure of the N-oxide group. From a chemical point of view, the N-oxide compounds differ from other compounds with the highest polarity (dipole moment of N-oxide of about 5 D, while the dipole moment of the source polyamine 0,65 D). This is due to their unique ability to absorb different is e toxic compounds, metals, etc. and then remove them from the body. It is a high adsorption capacity and polymeric nature due to expressed detoxification properties of these compounds.

Due to these properties, these compounds absorb various toxic substances, including metals, products of metabolism and other, and then remove them from the body. It is the adsorption capacity and the polymer due to the nature expressed detoxification and lipoic properties of these compounds.

In its detoxification properties of the copolymers in accordance with the invention is hundreds of times greater than in the detoxification properties of the known compounds of the same purposes, such as gemodez, albumin, dextran etc. however, the described copolymers have a noticeable immunomodulatory properties - stimulation coefficient varies in the range 3÷7.

When creating vaccinology and therapeutic preparations of new generation of high molecular weight immune carrier of antigens and allergens SP-NO provides greater stability of antigens, increased immunogenicity by reducing immunization doses of antigens, more efficient formation of immunological memory to antigens and increase preventive vaccine efficacy, as well as a high level of safety of the vaccine. Modificati the antigens of different nature of high molecular weight immune stimulator SP-NO allows you to enhance the immunogenic activity. The accession NP-NO to allergens leads to reduced allergenic activity, increasing the safety of the drug and reduce the risk of complications when introducing them in sensitized body. The binding of allergens with SP-NO provides an opportunity to significantly enhance the immune response, stimulate high level production of allergen-specific IgG antibodies that provide protection for Allergy sufferers when hit allergenspecific substances in his body.

As carriers of other pharmacologically active compounds JV-NO can be used by covalent binding, while covalent binding is achieved by the introduction of chemically highly reactive groups.

Evaluation of the immunogenic and protective properties of the obtained conjugates SP-NO with different antigens and including the hemagglutinin of influenza virus (HA), was carried out by studying the reactions of the immune system and the organism as a whole in experimental animals.

It has been found that the introduction of conjugate HA-SP-NO in the body of the animal it is induced by visible or antibody-based test response after a single dose of conjugate. In response to the reintroduction of developing a secondary, more intense response, the maximum is achieved after 10-15 days, he longer held in very high values even 3 months after the immunodeficiency is Itachi. In experiments on animals show high immunogenicity of the conjugate in the reaction of inhibition of haemagglutination of rtge. In the test block reproduction of the live virus in chicken embryos revealed a direct neutralizing activity of the resulting antibodies. For these tests, the conjugate is not inferior televizionnoi reference vaccine. In the analysis of isotypes of antibodies established that produces antibodies IgM, IgA, mainly IgG but not IgE. Initiated the formation of immune memory, which indicates the involvement of T cells in response to the introduction of the conjugate, is the accumulation of T-helper cells. What is very important is induced by intensive formation of virus-specific T-killers.

Derived SP-NO possess a wide spectrum of pharmacological activity, high bioavailability, capacity for excretion from the body, by the security application.

SP-NO in a wide range of doses activates macrophage link, humoral immune response, has anti-infective and anti-tumor activity, and increases the resistance of cell membranes to the cytotoxic effect and reduces the toxicity of drugs with a joint introduction.

SP-NO receive as a result of chemical synthesis by partial oxidation of the tertiary nitrogen in the main chain of the macromolecule source aliphatic polyamine containing nitrogen in the main chain in slightly acidic water, aqueous-alcoholic or alcoholic solutions. As the oxidizing agent can be hydrogen peroxide and other organic or inorganic peroxide or gidroperekisi, salts of oxygen-containing halogenation, ozone, or oxygen obtained by electrolysis of water.

The synthesis is carried out at a temperature of 20 ÷40°C. the Solvent and excess peroxide is removed by ultrafiltration, followed by freeze drying.

The identification and analysis of patterns obtained JV-NO and its derivatives is carried out using methods of physico-chemical analyses: NMR, PMR, IR-spectroscopy, UV-spectroscopy. The molecular-mass characteristics obtained using the chromatographic complex, which includes high-performance liquid chromatograph with 4 detectors - UV-spectrophotometry, refractometry, low-angle laser light scattering, fluorescence spectroscopy.

Upon receipt of the conjugates and complexes SP-NO with proteins and glycoproteins of different nature, having a primary amino group or carboxyl group, as the protein component of this reaction can be used:

- when receiving vaccinology drugs - antigen complexes secreted from the pathogen hepatitis a, b, C, tuberculosis, anthrax, cholera, typhoid, whooping cough, diphtheria, tetanus, Johann the x infections;

- upon receipt of drugs prolonged action with a high level of safety and efficiency - erythropoetin, cytokines, interferons, enzymes, cytostatics, antibiotics, hormones, and vitamins;

- upon receipt of funds specific immunotherapy - allergens and Allergology allocated from pollen and grasses, house dust, food, or other

In each case, vary the conditions of the reaction, the ratio of the components, pH, temperature of the reaction, the duration of the process, the concentration of reagents in the reaction mixture, which affects the yield and composition of the final product.

To use the SP-NO as a carrier of various pharmacologically active compounds can be applied to methods of binding the complexation due to multipoint electrostatic interactions of the components or covalent binding.

If the antigen - individual compounds with known chemical structure and structure - VI-antigen, hemagglutinin, enzymes, etc., it is advisable to choose covalent binding. If it is a mixture of different substances, it is preferable to bind complexation reactions, including the use of polymer-antigen-metal complexes.

In the case of oppositely charged compounds complexor the Finance in certain circumstances occurs with high speed and output. In the case of the same charged components of the preferred methods of obtaining with the formation of the polymer-antigen-metal complexes. For example, in the specific case of using as allergoids pollen of Timothy grass or birch, which is not an individual connection. When linking allergoids with SP-NO both compounds have a weak negative charge, i.e. charged the same name. In this case, choose the complexation of using the polymer-metal complexes.

The basic criteria are safety, efficiency and adaptability of processes for the synthesis of compounds and receiving pharmacological preparations.

The comparison of different methods of binding revealed the benefits of it education ternary polymer-antigen-metal complexes. Manufacturability extremely high - on the background of preliminary complexation of two compounds, add the calculated amount of salt of copper with the formation of the "staples" between the molecules, which leads to the stability of the complex under physiological conditions.

The formation of ternary polymer-metal complexes based on the SP-NO, and ions of transition metals and antigens or allergens was one of the best ways in which a sufficiently strong binding of the antigen and polymer wear what I not accompanied by changes in their chemical structure and, as a consequence, the spectrum of biological action of components, does not produce toxic by-products of the reaction and has a number of technological advantages as a method of obtaining a ternary polymer-metal complexes is simple, technological, proceeds in one stage under mild conditions and with 100% yield.

In the generated vaccine or pharmaceutical compositions immunomodulatory, anti-infective and anti-inflammatory effect is due to a combination of physiological activity JV-with NO active components.

The following examples illustrate the possibility of obtaining copolymers in accordance with the present invention and the results of the study of their properties

In the following examples 1-5 are examples of synthesis of specific SP-NO; examples 6-13 illustrate the study of the pharmacological properties of the SP-NO; in examples 14-20 presents the possibility of obtaining conjugates and complexes derived SP-NO different physiologically active components, leading to the formation of vaccine and pharmaceutical compositions based on them.

In examples 1-3 show variants of synthesis of JV-NO when using three source hetero-chain aliphatic polyamines, respectively, poly-1,2-conidine SP-NO-1, poly-1,4-Hinkley CII-NO-2, poly-1,4-triethylenediamine SP-NO-3.

Example 1. Synthesis of copolymer of conidi the a and N-oxide conidia (SP-NO-1-1).

For the synthesis of the copolymer used veteriany aliphatic polyamine poly-1,2-Konigin.

10 g of the original polyamine with 70000 MM D dissolved in 300 ml of 96%ethyl alcohol. The solution is cooled to a temperature of 4÷6°C. and while stirring, add 20 ml of 30%hydrogen peroxide. The oxidation reaction polyamine carried out for 10 hours, after which part of the solvent is removed in vacuum and add 300 ml of water. Removing unreacted hydrogen peroxide is carried out using an ultrafiltration installation "Pelican". The resulting solution of the copolymer conidine and N-oxide conidine concentrated and dried on lyophilization installation. After drying the yield of the target product is 99%, n=620, q=0,35n, z=0,65n.

Elemental analysis data:

Calculated, %: C To 66.3; H 10,24; N11,20.

Received %To 66.4; H 10,31; N 11,22.

The characteristic band of the compound obtained in the result of analysis by the method of IR-spectrometry

is 960 cm-1and 1130 cm-1.

Example 2. Synthesis of copolymer of 1,4-Hinkley and N-oxide 1,4-hinoklidina (SP-NO-2-1).

Poly-1,4-Hinkley with 75000 MM D 10 g dissolved in 150 ml 0,1N acetic acid. When cooled (4°C) and stirring, add 10 ml of 30%hydrogen peroxide. The solution is maintained under these conditions for 24 hours. Then the solution is subjected to ultrafiltration to remove excess hydrogen peroxide is acetic acid. An aqueous solution of the copolymer is dried by lyophilization.

The characteristic band of the compound obtained in the result of analysis by the method of IR-spectrometry, is 960 cm-1and 1130 cm-1.

The product yield 100%: n=650, q=0,2n, z=0,8n.

Example 3. Synthesis of copolymer of 1,4-triethylenediamine and N-oxide of triethylenediamine (SP-NO-3-1).

10 g of the original polyamine poly-1,4-triethylenediamine with MM 80000 D dissolved in 400 ml of acetic acid at pH 5. After dissolution polyamine under stirring and cooling (4-6°C) add 20 ml of 30%hydrogen peroxide. The reaction is carried out within 35 hours. After completion of the reaction, the obtained solution was purified from excess unreacted components using the method of ultrafiltration treatment and then subjected to freeze-drying. After drying the yield of the target product with the following characteristics: n=700, q=0,25n, z=0,75n is 100%.

Table 1 shows the characteristics (n, q, z and MM) copolymers conidine and N-oxide conidia (SP-NO-1-1, SP-NO-1-2, SP-NO-1-3), copolymers of 1,4-Hinkley and N-oxide 1,4-hinoklidina (SP-NO-2-1, SP-NO-2-2, SP-NO-2-3) and copolymers of 1,4-triethylenediamine and N-oxide of triethylenediamine (SP-NO-3-1, SP-NO-3-2, SP-NO-3-3)obtained according to the methods described in the examples, respectively, 1, 2, and 3 with varying ratios of the starting components and reaction parameters.

Example 4. Study of the pharmacokinetics of R is slichnih types SP-NO, having a radioactive tag With13.

Investigation of the pharmacokinetics of labeled SP-MO performed by standard methods in intramuscular dosage form JV-NO in a dose of 20 mg/kg 0.75 in (MBq/kg) to rats.

Observations of the animals showed no tissue comunali of the drug. Found heterogeneity in the distribution of SP-NO in organs and tissues in males and females. Excretion SP-NO occurs mainly in two phases. The half-life of the rapid phase - 1.5 hours, slow - 84 hours.

The results of pharmacokinetic studies SP-NO-1-1, SP-NO-2-1 and Cn-NO-3-1, presented in table 2 show that SP-NO is rapidly absorbed into the systemic circulation and reaches its maximum concentration after 30-50 min Period poluraspredelenia about 0.5 hour period preliminaly - 20-46 hours, and the average residence time of drug in the body is about 40 hours.

Example 5. Study of antioxidant properties of SP-NO.

The ability of SP-NO suppress the formation of reactive oxygen species (ROS) is estimated in the system of interaction of hydrogen peroxide with horseradish peroxidase (Reanal). When this registration education superoxide anion radicals is performed using chemiluminescence intensity of oxidation lyuminola products specified reaction. The chemiluminescence analysis performed on a 36-channel installation "Luficer-B is at a temperature of 37°C.

Pre-prepare a collaborative environment consisting of: saline, phosphate buffered (pH 7.2÷7,4), luminal (Sigma Chemikal Co. at a final concentration of 0.6×10-1M) and interacting reagents hydrogen peroxide (at a concentration of up to 0.005% and horseradish peroxidase (final concentration 1 μg/ml).

The specified ratio of the reagents provides for the preservation of intense chemiluminescence level 300000 pulse/sec for 40 minutes. The mixture is placed in a volume of 500 ál in test tubes of chemiluminometer and record levels of illumination for 5 minutes Then in the appropriate tube type analyte (SP-NO-1-1, SP-NO-2-1, SP-NO-3-1, SP-NO-3-2) in a volume of 10 µl. The range of investigated concentrations of 25 to 250 mcg/ml During this procedure was dose-dependent damping signal for 10-90% of the original level. Measurement stop on the moving curve chemiluminescence plateau (15-20 minutes). For each concentration of the deposited substance calculate the area under the curve and compared it with that of the control tubes with saline.

The results are expressed as the percent suppression of free radical reactions for each concentration of the investigated substances. For comparison, the antiradical activity of different samples calculate the concentration at which is suppressed and the efficiency to 50% of free radicals.

The data presented in graphs figure 1, indicate that the add in the radical reactions of all the samples SP-NO at a concentration of 15 μg/ml leads to the suppression of the level of radical reactions to a much greater extent (50%) in comparison with the known compound (reference drug) in the same dose (30%), which confirms the high antioxidant properties of these compounds.

Example 6. The study of detoxification properties SP-NO.

Evaluation of the protective properties of SP-NO spend on models of acute toxicity.

In studies using white mice - hybrids (model CBA×C57BL/6F1) weighing 18-20 g

Prepare 0,04%solutions used in experimental studies to be toxic to mice substances of natural origin - MRM with addition and without SP-NO in the ratio of 1:1 and 1:5. The compounds administered to the animals once intraperitoneally in a volume of 0.25 and 0.5 ml doses, respectively. Animals of the control group injected with 0,5ml of saline. Observations of the animals should be performed within 21 days.

The data presented in table 3, illustrate the protective properties of the SP-NO.

Example 7. The study of antidote properties SP-NO.

Studies conducted in vivo in acute poisoning CuSO4. The drugs enter the 7 groups of animals, among them animals of the first group type Fitr is the target, animals 2-4 groups only CuSO4in lethal to animals doses (mg/kg): 12,5; and 25,0 50,0, and animal 5-7 groups administered similar doses CuSO4in combination with similar doses of SP-NO-3-1.

The research results presented in table 4 indicate pronounced is the effect of SP-NO.

It was found that the simultaneous introduction of the SP-NO-3-1 dose of 12.5 mg/kg and above three lethal doses CuSO4not only protects animals from death, but also prevents the development of symptoms of intoxication.

Example 8. Evaluation of the protective properties of SP-NO model of hemolysis of erythrocytes.

Studies were performed on quartz (SiO2) hemolysis of erythrocytes using international standard quartz dust DQ-12 particle size less than 3 μm according to the method A.David (1976). Human blood was taken with heparin, erythrocytes washed three times in 10-fold volume of colorless Hanks solution, followed by centrifugation at 3000 rpm for 10 minutes Erythrocyte suspension was respondible to 4%concentration (10 cells in 1 ml). All studies were performed in 3-5 parallel samples. Samples of SiO2and red blood cells were selected with constant stirring on a magnetic stirrer.

1 ml of a 4%suspension of erythrocytes were mixed with 1 ml of the study drug, was added 1 ml of 0.3% (3 mg/ml) suspension SiO2and incubated in the tech is of one hour at 37°C, gently shaking the sample every 5 minutes. After incubation, all samples were added to 7 ml of buffer, centrifuged and the supernatant was photometrically on SF-26 at 540 nm. The results of the hemolysis was expressed in percent, with 100% of the hemoglobin content in samples of 1 ml of a 4%suspension of erythrocytes with 9 ml of distilled water. The protective effect of the study drug was calculated by the formula and expressed as a percentage:

where

E540For sio2- extinction test specimen in the test tube water-silicon-erythrocytes";

E540experience the extinction of prototypes in vitro drug in the buffer silicon erythrocytes".

Presented in table 5, the data show that the silicon dioxide has pronounced hemolytic properties. 3 mg SiO2within hours of incubation causes hemolysis 87.4% of erythrocytes. Incubation of erythrocytes with SiO2in the presence of 50 µg SP-NO-3-2 almost completely protects erythrocytes from hemolysis.

Property JV-NO to protect erythrocytes from hemolysis compared with high molecular plazmozameschayuschie solutions of polyvinylpyrrolidone (gemodez), dextran (poliglyukin) and albumin. As the buffer used colorless Hanks solution.

Polyvinylpyrrolidone and albumin also have protecting properties, but the dramatic effect is istihaada only at high doses. When this drug will not destroy, can accumulate in the body and cause unwanted side effects.

The data presented in table 5 clearly indicate the expressed properties SP-NO as a detoxifier and their advantages in comparison with recognized detoxicate.

Example 9. The study of the immunomodulatory activity of SP-NO

Together with immunomodulating activity of SP-NO was assessed by their ability to stimulate antibody production to sheep red blood cells or antigens protein nature (In the-subunit of the cholera toxin, tetanus toxoid) in experimental mice-the first generation hybrids obtained by crossing mice lines CBA and C57BL (model CBA×C57BL/6F1) by determining the number of antibody productive cells (AFC) in the spleen of mice in 4 to 7 days after their introduction. The experiment according to the conventional method EPHE. The number of AFC in the spleen of mice determined by the method of local hemolysis agar. The titers of antibodies generated in response to the introduction of protein antigens, determined by the method of enzyme immunoassay. Together with immunomodulating efficiency JV-NO estimate on the ratio of the number of the KLA, formed by the joint introduction of antigens and SP-NO among the KLA in the group of control animals. Doses range from 1 to 1000 mg/kg mouse intraperitoneal and subcutaneous injection.

p> Table 6 shows the values of coefficient of stimulation depending on the specific compounds and dose (dose ranged from 1 to 1000 mg/mouse, intraperitoneal and subcutaneous injection). The index of stimulation of the immune response has a value of from 3 to 7, i.e. comparable with the known standard toxic stimulant - polyacrylic acid (PAC). Experiments have shown that there is a dependence of the coefficient of stimulation on the dose and on the composition of SP-NO.

The results of the study indicate a high immunomodulatory activity of SP-NO, which is manifested in a wide range of doses with different ways of introduction.

Example 10. Receiving anti-TB vaccines through complexation antigenic complex, allocated from titanoboa extract of mycobacteria BCG with SP-NO. Evaluation of the protective properties of the drug in the experiment on animals.

Obtained and examined a number of samples of TB wakciniruemogo drug with optimized ratio of antigen and SP-NO-3-1.

As antigen was selected antigenic complex (AC), consisting of glycopeptides allocated from titanoboa extract the cell walls of mycobacteria BCG.

According to the passport data on antigen protein content of 1 mg of dry matter AK account for 50.2 ág. Determination of protein avodat according to Bradford.

Upon receipt of sample wakciniruemogo preparation proceeded from the estimated doses for antigen - 100 µg and 50 µg AK on mouse dose of protein, respectively, was 5 μg and 2.5 μg per mouse. Dose of SP-NO-3-1 was selected on the basis of the previously obtained results and was 1000 μg per mouse. To obtain vaccinology drugs used connection SP-NO-3-1. Determining the components of the drug calculations were made on the content of the basic substance of the SP-NO-3-1.

The reaction of complex formation is as follows.

500 mg SP-NO-3-1 was dissolved in 10 ml of 0.05 M phosphate buffer pH 5.8. At a temperature of 2-4°C add 100 ml of solution AK in 2 ml of the same buffer. When the pH of the macromolecule polymer carrier and antigens are charged oppositely.

The complexation of conduct by the slow introduction of a solution of the antigen in a solution of JV-NO-3-1, preventing sedimentation. Increasing the solubility of the antigenic complex in water exercise due to its complexation with SP-NO. In one of the options (Drug 1) add a solution of the pharmaceutical composition SP-NO to antigenic complex with constant stirring and pH control. In another embodiment (Product 2) conduct consolidation solutions antigen and SP-NO with constant stirring, cooling and pH control.

After conducting the Oia synthesis solutions of drugs subjected to freeze drying and control.

Determination of protein content and the analysis is performed using fluorescence spectroscopy and polyacrylamide gel electrophoresis (SDS page). The characteristics of the two preparations obtained by the described method are shown in table 7.

Protective activity of the synthesized drugs 1 and 2 are assessed by two indicators - Visavuori bacilli from the lungs and spleen of infected mice, as well as in terms of the survival of animals after infection.

For immunization of mice used 3 preparation:

The drug 1 contains 1 dose of 5 μg of antigen and 500 mcg SP-NO;

Drug 2 contains 1 dose of 2.5 μg of antigen and 500 mcg SP-NO;

Preparation 3 contains 1 dose of 500 µg SP-NO.

Mice subjected to immunization preparations 1, 2 and 3 subcutaneously at 1 point in a volume of 0.2 ml twice with an interval in 2 weeks. As a positive control, use of animals that received 1 injection of BCG (Prague 106). As a negative control use unvaccinated animals.

5 weeks after the second immunization of experimental animals of all groups infect a lethal dose of Mycobacterium tuberculosis H37Rv (5×106KOE).

The inoculation of bacilli from the authorities determined 3 weeks after administration of lethal doses of mycobacteria. According to the results shown in table 8, there is a decrease in the number of CFU in the lungs and spleen of the mice vaccinated with the drug and, compared with the number of CFU in 4 control group of animals received an injection of lethal doses of bacteria.

On the survival of experimental animals, the most effective is the Drug 2. This drug exerts a protective effect against lethal doses of mycobacteria in 57% of the animals to the 70th day of observation, but the drug was somewhat less effective than BCG. Double introduction animal Drugs 1 and 2 to 16 days lengthens the lifetime of lethally infected mice.

The data presented in table 9, show the effectiveness of the obtained vaccine on survival.

Thus, according to the data obtained the greatest protective effect possessed the drug, in which the protein content (in the AK) and SP-NO, respectively 5 μg and 500 μg. Its effect was significantly higher than that in the case of other used drugs.

Example 11. Obtaining conjugates SP-NO with proteins and glycoproteins of different nature, having a primary amino group in the side chain.

Upon receipt of the conjugates and complexes SP-NO with proteins and glycoproteins of different nature, having a primary amino group or carboxyl group, as the protein component of this reaction can be used:

- when receiving vaccinology drugs - antigen complexes secreted from the pathogen hepatitis a, b, C, TB is uleta, anthrax, cholera, typhoid, whooping cough, diphtheria, tetanus, pyogenic infections;

- upon receipt of drugs prolonged action with a high level of safety and efficiency - erythropoetin, cytokines, interferons, enzymes, cytostatics, antibiotics, hormones, and vitamins;

- upon receipt of funds specific immunotherapy - allergens and Allergology allocated from pollen and grasses, house dust, food, or other

In each case, vary the conditions of the reaction, the ratio of the components, pH, temperature of the reaction, the duration of the process, the concentration of reagents in the reaction mixture, which affects the yield and composition of the final product.

Method of obtaining conjugates SP-NO protein in General has the following procedure: 10 mg of SP-NO, obtained according to the above examples, dissolved in 1 ml of water. To the solution was added 5 mg bromoacetonitrile and the mixture is stirred for 3 hours at 30°C. Then the reaction mixture was added a solution of 3 mg of protein in 0,01M phosphate buffer pH of 8.2. The conjugation reaction is carried out at a temperature of 2-4°C for 20 hours. After completion of the reaction, the reaction mixture was purified by ultrafiltration, the target product emit lyophilic drying.

Por what measures 12. Obtaining conjugates SP-NO with polysaccharide antigens of nature (PS-AG) to obtain vaccinology compounds with enhanced immunogenicity and protective effect against infections.

Obtaining conjugates SP-NO with polysaccharide antigens of nature (PS-AG) is shown in example VI capsular-polysaccharide having a protective activity against the infection caused by Salmonella typhi.

Conjugates PS-AG JV-NO gain according to the method described in example 11, by formation of a covalent bond between the carboxyl group of residue N,O-acetylgalactosamine acid PS-AG and hydrazide group of activated derivatives of SP-NO, using a condensing agent, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). The condensation reaction of PS-AG with a hydrazide derivative of polyoxidonium SP-NO is carried out in water-salt solutions (0.2-0.35 M NaCl) at a weight ratio between the components equal to 1:0.25 to 1:10, taken at a concentration of 0.1-0.3% for PS-AG and 0.07-1% NP-NO for 3-20 hours at a temperature of 4-20°C and a pH of 5.6 to 5.8; the amount of EDC was 0.2-0.5 wt %.

The procedure for allocation of the conjugate from the reaction mixture carried out on a column (of 2.6×70 cm) media Sephacryl S-1000 (Pharmacia, Sweden) in 0.2 M NaCl solution. The fraction with a conjugate that go with the free volume of the column are pooled, concentrated in vacuo to a small volume, dial the form against physiological saline, containing 0.01% of thimerosal as a preservative, and stored at 3-8°C.

Output conjugates, calculated on the PS-AG, 60=10% of the original weight of the number of PS-AG.

Processing conjugate solution of 0.5 M NaCl solution or 0.25% solution of the detergent deoxycholate sodium does not reduce the molecular weight of the obtained product according to gel-chromatography on Sephacryl S-1000 and Sepharose 4B, indicating the presence of covalent bonds between the PS-AG and SP-NO.

The presence of PS-AG in the resulting conjugates confirm serological analysis. Conjugates inhibit the Treponema pallidum haemagglutination assay with anoreceptive anticorodal at a concentration of 0.12 to 250 µg/ml of Conjugates of PS-AG for the single intraperitoneal immunization of laboratory animals (mouse lines F1) at a dose of 5-100 μg/mouse produce specific serum antibodies in number, are 2-4 times larger than the original PS-AG.

An example of a sample No. 1 - conjugated VI-polysaccharide (VI-PS) with a molecular mass of 3-5 MD, representing a linear homopolymer consisting of residues 3-O-acetyl-2-acetamido-2-desexed-the galacturonic acid linked (1-4)-glycosidic bonds, with a hydrazide derivative of SP-NO, synthesized by the method of example 5. Obtaining a conjugated capsular VI-polysaccharide at a mass ratio of VI-SS: SP-NO 1:0,2.

To 10 mg VI-PS 1.7 ml of 0.2 M NaCl solution at pH of 5.6 was added with stirring 2.5 mg SP-NO 1.7 ml of 0.2 M NaCl solution at pH of 5.6. The resulting solution was stirred for 30 minutes at 20°C, add 8 mg dry EDC. pH of 5.6 to 5.8, support 0.1 N. HCl solution for 3 hours at 20°C and 17 hours at 4°C.

The reaction mixture cialiswhat during the day in dialysis tubes (6000-8000, Spectrapro, USA) against 0.2 M NaCl solution. Dialysate applied on the column with the carrier Sephacryl S-1000 (2,6×70 cm). The elution carried out with 0.2 M NaCl at 60 ml/hour. The registration is carried out spectrophotometrically at 206 nm and refractometrically.

Fractions corresponding to the free volume of the column, K=0,1-0,36 are pooled, evaporated to a small volume on a rotary film evaporator, dialist against saline with 0.01% of thimerosal pH 7.0 and stored at 3-8°C. the Yield of conjugate VI-PS is 70%, the mass ratio between the components of the VI-TS:SP-NO in the conjugate is 1:0,3.

An example of a sample No. 2 is conjugated to hydrazide derivative SP-NO, synthesized according to example 6 VI capsular-polysaccharide at a mass ratio of VI-SS:SP-NO 1:0,5.

Unlike obtain sample No. 1 to 10 mg VI-PS in 3 ml of 0.2 M NaCl solution under stirring and pH of 5.6 add 5 mg of SP-NO in 3 ml of 0.2 M solution of NaCl at a pH of 5.6. Dry the EDC was added to the share of the ve 12 mg.

The output of the conjugate VI-PS is 70%, the mass ratio between the components of the VI-TS: SP-NO in the conjugate is 1:0,3.

Example No. 13. The study of protective properties of conjugates SP-NO PS-AG as vaccinology compounds with enhanced immunogenicity and protective effect against infections caused by pathogens Salmonella.

Studies conducted on the vaccine samples No. 1 and No. 2 obtained in accordance with the technology described in example 15, in comparison with native VI-polysaccharide (VI-PS).

Drugs tested on laboratory animals the mice CBAxC57B1/6)F1 tests in active protection of mice from infection control animals production virulent strain of typhoid fever Salmonella typhi Ty2 No. 4446. Test active protection of mice is the main method for laboratory evaluation of effectiveness bruchnotifozhnykh vaccines. The requirement for vaccine preparation, studied in this test, the protection of mice after infection the culture at a dose of 3-6 LD50. VI-PS and immunochemical active compared with SP-NO samples # 1 and # 2 enter only once. Use intraperitoneal immunization and infection, as this method of introduction has the most stringent requirements for immunogenicity of the vaccine. Contamination spend a month or more after the introduction of the experimental vaccine.

Comparison of the protective effect of series con who Ugarov samples No. 1 and No. 2 and commercial bruchnotifosny vaccine. Use the vaccine bruchnotifosny chemical sorbirovannoe liquid. Drugs give is equal to the VI antigen dose.

Comparison of the protective properties also hold for the model with infection with the virulent strain. Use bruchnotifosny chemical sorbed liquid vaccine that is recommended for mass immunization in the Russian Federation. Drugs give equal Vi-antigen dose.

Data on survival of animals are presented in table 10, for samples No. 1 and No. 2 and compared with a commercial vaccine show that the conjugation VI-PS antigen with SP-MO allows you to significantly increase its vaccinifolia properties and provide 100% protection of experimental animals during infection dose, strain not less than 3 LD50.

The binding of PS-AG with a synthetic polymer immunomodulator SP-NO allows you to get the drug with increased immunogenicity.

Example No. 14. The study of progenote series of sample No. 1 and the commercial vaccine.

Experiments conducted on 9 rabbits of the chinchilla breed, which enter the sample No. 1 in a dose of 1000 μg and 200 μg and commercial vaccine at a dose of 40 mg. thermometry results presented in table 11, indicate that the pyrogenic dose (1000 µg) vaccine VI-PS JV-NO significantly exceeds pirogennoe dose commercial bruchnotifosny vaccine (40 µg).

The results of studies contained in the application is Oh 16-18, show that the conjugates of the polysaccharide VI antigen and SP-NO to use against typhoid significantly improve protective immunity while reducing progenote drug.

The binding of SP-AG with a synthetic polymer immunomodulator SP-NO allows you to get immunochemical active drug with increased immunogenicity, reduced pyrogenalum and can be recommended for developing a vaccine preparation against bruchnotifosny infection.

Example 15. Using the SP-NO to obtain a conjugate with the medicinal substance.

Shown at the example of a conjugate SP-NO with lidsay (enzyme hyaluronidase).

Getting conjugate SP-NO drug substance can be carried out, for example, by the condensation reaction:

100 mg hydrazide SP-NO dissolved in 4 ml of 1 N. HCl. The solution is cooled to 2-5°C. Then, while stirring and cooling add to 1.15 ml of 3% solution of sodium nitrite. After 15 minutes, bring the pH to 8.5 by adding 2 N. NaOH.

A solution of 20 mg of enzyme in 10 ml of 0.05 M phosphate buffer pH 8.5 added to a solution of hydrazide SP-NO. Maintain the pH of the reaction mixture equal to 8.5 by adding 2 N. NaOH. The reaction is carried out for 12 hours under stirring and cooling (0-2°C). For isolation and purification of the conjugate reaction mixture is applied on a column (of 2.6×90 cm), sepolgen the th Biogel P-100, as eluent using phosphate buffer 0,05M pH 7.5, containing 0,05M NaCl.

The output of the conjugate is controlled using a flow-through spectrophotometer at 226 nm. Determination of protein content and analysis of the conjugate is carried out using fluorescence spectroscopy, electrophoresis in SDS page. 1 mg product contains 0.2 mg of enzyme.

Example 16. The study protivovirusnyh properties of the drug based on the conjugate SP-NO and lidz (Product L), obtained according to example 15.

A study conducted on rats male Wistar rats with initial weight of 180-200 g model pneumovirus caused odnokratno intratracheal introduction of 20 mg of silica dust. The effectiveness of the Drug L was estimated by the content of the main components of connective tissue, which adequately reflect the degree of fibrosis. In the lungs was determined by the content of lipids, collagen proteins, glycoproteins, glycosoaminoglycans, was also studied histomorphological changes and ultrastructure of the lungs.

One group of animals the Drug L was administered 4 days after dusting (prophylactic use). Another group after 1 month, i.e. on the background developed fibrotic process in the lungs. The drug L was administered intraperitoneally at a dose of 1500 ME 1 time a week for 1 month. The third group of animals received native hyaluronidase, having the same total activity.

LM is now scored in 1, 2 and 3 months. It is established that native hyaluronidase no effect on the fibrotic process in the lungs. At the same time, the Drug L not only delays the development pneumovirinae process, but also possesses a strong ability to dissolve existing in the lungs fibrous tissue.

Experimental studies on the model of pneumoconiosis (silicosis) demonstrates the ability to effectively impact on the fibrotic process through Preparation of L. find the optimal pharmacological dose and dosage regimen of the Drug L at which the drug is not only delays the further development of pneumovirus, but it brings back the development of granulomas in the lungs, which is confirmed by biochemical, histological and electron microscopic studies. For rats the best results to contain the growth of connective tissue destruction and fibrosis are able to achieve with the Drug L-1 per week dose of 500 IU/kg, that is equal to 7 EUR/kg in terms of activity Lidz. Lydasum at this dose and effect has not had any noticeable positive effect on the course of fibrosis in the lung.

In the implementation of therapeutic action of the Drug L play a decisive role in physico-chemical and, as a consequence, the pharmacological properties inherent in the media SP-NO. Reverse the development of fibrous tissue under the influence of the Drug L suggests, that the drug has the ability not only to cause degradation of the connective tissue, but also to prevent the degradation products and released from silicotics knots particles of silicon dioxide to re-stimulate the fibrotic process.

This example illustrates the ability to create medicines with new medicinal properties by conjugation JV-with NO known medicinal substance.

Example 17. Using the SP-NO as immunostimulating component upon receipt of the products for specific immunotherapy.

Specific immunotherapy or desensitization is glavnom etiological treatment of patients with allergic diseases.

This method gives in 70-80% of patients with good sustainable results, not without some drawbacks that limit its applicability. The disadvantages are: 1) the risk of allergic reactions during the course of immunotherapy, both local and system; 2) insufficient high immunogenic activity of small doses of specific allergenic preparations to initiate the biosynthesis of blocking IgG-AT. Therefore, one of the main tasks of Allergy is to develop methods of modifying allergenic drugs to reduce their allergenic activity and increased immunogenicity./p>

One of the techniques of modification of the allergen is the binding of allergenic and allergenic drugs with high molecular Immunostimulants that may enhance their immunogenicity, to foster a high level of production of allergen-specific IgG antibodies and reduce the possibility of complications hypoinsulinaemia therapy.

Modified derivatives JV-have NO reactivity, allowing covalently or complex to bind molecules of different nature, having any functional group.

The choice of tactics of binding depends on the nature of the antigenic component. If the antigen - individual compounds with known chemical structure and structure - VI-antigen, hemagglutinin, enzymes, etc. may conduct covalent binding. If it is a mixture of various substances - complexation in the case of oppositely charged compounds and ternary polymer-antigen-metal complexes in the case of like charged molecules.

The possibility of obtaining Allergology, antigenic component which is a mixture of various substances, shown at the example of a prototype of Allergology based allergoids pollen of Timothy grass or birch and SP-NO.

To obtain prototypes of Allergology based allergoids pollen of Timothy grass or birch and SP-N was chosen as the method of formation of ternary polymer-metal complexes.

Security is provided by the presence of only 5 of copper ions at 600 weakly charged N-oxide groups. Manufacturability is extremely high - in the background of pre-complexation of the two compounds is added to the estimated amount of the salt of copper with the formation of the "staples" between the molecules, which leads to the stability of the complex under physiological conditions.

Example 18. Getting Allergology based allergoids (ALD) pollen of Timothy grass or birch and SP-NO (on the example of CNO-3-1).

5 mg CNO dissolved in 10 ml of 0, 1 N phosphate buffer pH 7,2 add 5,0 µg CuSO4.

Then, with vigorous stirring, 5,0 µg allergoids pollen of Timothy grass (or birch), dissolved in 5 ml of phosphate buffer. The mixture was incubated for 8 hours at a temperature of 4±2°C. the Solution is freeze-dried. The resulting preparation is the ratio of ALD: SP-NO is 1:100.

Changing the ratio of components in the reaction mixture get drugs with different amounts of protein and various proportions of Allergoids and SP-NO in the preparations. For assessment of allergenic activity selected three sample obtained according to the described method, with the contents of allergoids (determination of protein by the method of kildala) in the composition of the conjugates:

in sample 1 of ALD: SP-NO 1:100, 10 μg protein/mg of the drug;

in sample 2 And: SP-NO - 1:20, 50 µg protein/mg;

<> in sample 3 And: SP-NO - 1:10, protein 41 mcg/mg.

Example 19. Assessment of the allergenic activity of the complex products in vivo. The definition of immunogenic activity of various forms of allergenic preparations.

Assessment of the allergenic activity of the complex products in vivo was carried out using methods of active cutaneous anaphylaxis (AKA) and passive cutaneous anaphylaxis (PKA) in sensitized Guinea pigs, and also for studying the ability of the conjugated forms of medication can initiate the formation of IgE-AT mice. These methods allow to adequately assess the allergenic properties of drugs.

Test of inhibition when performing enzyme-linked immunosorbent assay (ELISA) showed that conjugated drugs reduce the allergenic activity of native products by 60%.

The study of the allergenic activity of the native allergen, allergoids and samples No. 1, No. 2, No. 3 in the reaction AKA sensitized Guinea pigs.

Sensitization of Guinea pigs allergen of birch formed the sustainable hypersensitivity both immediate and late types.

The intensity of allergic reactions to a specific allergen is practically unchanged since 3 weeks after the onset of sensitization over the next three months.

Sensitization is expressed in the presence of positive reactions AKA, the intensity of which is avisit the number entered intradermal native allergen. Dose dependence of the intensity of the reaction AKA evident as when registering diameter of the colored spots on the injection of allergen, so when determining the number of extracted colors.

The dose of the injected allergen 5 µg reaction AKA evolved only in individual animals. On the other hand, the increase in the number of intradermally injected allergen over 100 µg caused coverage not only of the skin, and underlying tissue, making it difficult to assess the reaction. Based on this, mainly the number of trials used doses of allergen 5-100 mcg.

With the introduction of allergoids and sample No. 2 in such doses was significantly weaker than the corresponding dose of native allergen, samples No. 1 and No. 3. The intensity of the AKA, which is determined by the amount of released dye in place of an allergic reaction, with the introduction of allergoids and sample No. 2 was reduced compared to the native control, samples No. 1 and No. 3, when using 100 μg of allergen by 25%, with the introduction of 50 μg - 30% with the introduction of 25 μg - 10%. With the introduction of 12.5 µg most animals immediate allergic reaction has not been developed.

Allergenic activity of the complex products in the reaction PKA

Sensitization of mice with allergen, allergoids and wakciniruemogo drugs samples No. 1, No. 2 and No. 3 causes the formation of IgG-AT, which determine when post the information PKA rats in dilutions 2/2-1/32 with the introduction of the resolving dose of native allergen with Evans blue.

Maximum titer of serum, obtained by sensitization of mice native allergen and sample No. 1, was equal to 1/16. Options allergoids and samples No. 2 and No. 3 received the titer of the serum was significantly reduced. Thus, using reactions AKA and PKA shown that conjugated of ALD with SP-NO in the ratio of 1:20 and 1:10 reduces the formation of IgG-AT in the blood of sensitized animals.

The effect of immunization with allergen of birch, allergoids and complex preparations of ALD with SP-NO education IgG-AT

In the immunization of mice with native allergen, allergoids and wakciniruemogo drugs samples No. 1, No. 2 and No. 3 after 3, 5, 6, 7 weeks after the beginning of immunization in animals in the blood was determined IgG-AT in the credits 1/8-1/32.

The level of IgG-AT the immunized mice in all varianto experience has changed over time. Thus, the titer of the serum to the 21 day options allergoids, allergen, sample No. 1 was equal to 1/32, and when immunization samples No. 2 and No. 3, it reached the maximum value only on day 42 after the start of immunization and remained with them at a high enough level to 49 days. The titer of the serum, which was determined by the level of IgG-AT variant with allergoids, declined until 42 days and slightly increased to 49 days from the start of immunization.

From the data obtained it follows that the allergens in the form of complexes with SP-NO cause more in insignia immune response.

We studied the efficiency of conjugation of ALD with SP-NO method specific hyposensitization sensitized Guinea pigs.

Upon completion of the course specific hypersensibility intensity AKA was the most in control animals, it is taken for 100%.

The Hyper-sensibilization of the native allergen and sample No. 1 reduced the intensity AKA, compared with control, with the introduction of 200 micrograms of allergen 35% with the introduction of 25 micrograms of allergen by 50%. Much more effective was the Hyper-sensibilization of the sample No. 2. Compared to control the intensity of the AKA is reduced in this case, with the introduction of 200 micrograms of allergen 65%and with the introduction of 25 micrograms of allergen almost 100%.

The intensity of the response to allergoids declined with the introduction of 200 micrograms of allergen birch 50%.

Similar results were obtained when hypersensibility Guinea pigs of different forms of allergens in the reactions of PKA.

The maximum titer of serum 1/32 obtained in all variants of studies. However, the amount of released dye from the skin of animals, which corresponds to the intensity of reaction, in the experiments with sample No. 1 was observed (50%) compared with the reaction in the animals which were injected sample No. 2 and allergoids.

Using ELISA in serum giposensibilization Guinea pigs of different forms of allergens from pollen b the cuts determine the level of allergen-specific IgG-AT.

The research results show that the titers of IgG-AT animals, giposensibilization allergoids and sample # 1 is 1/8, which is higher than the control, whereas hyposensitization sample No. 2, the titer of IgG-AT is equal to 1/6. Thus, of all the drugs studied most high immunogenic activity has sample No. 2 in the injection of 5.0 ug.

As a result, studies have shown:

1. Chelation allergens reduces the risk of anaphylactic-type reactions.

2. The introduction of allergens in combination with SP-NO in the body of the animal is not accompanied by an increase in IgE antibodies in the blood.

3. Allergens in combination with SP-NO when introduced into the animal organism stimulate the increase of IgG-antibodies.

4. Drugs allergen, allergoids and Old: SP-NO in the test doses (250 μg protein / kg of body weight is alive.) non-toxic.

5. Complex allergic - SP-NO (sample No. 1) has (in comparison with other drugs) is more pronounced (1.5-2 times) hypoinsulinaemia activity.

6. Complexation of allergens can be regarded as a General methodological principle, possible to achieve the reduction of the risk of anaphylaxis to the introduction of allergens into the body.

In example 20 describes the obtaining of vaccine compositions based on the SP-as NO substances with immunoadjuvant action. Using patent-pending connection the settings of the JV-NO-1-1, SP-NO-2-1, SP-NO-3-1 to obtain the vaccine compositions due to their pronounced immunoadjuvant properties.

Example 20. Getting the vaccine compositions of the example of combined adjuvant Giacconi against hepatitis a and B.

Combined hepatitis di-(a+b)-a vaccine with the use of patented compounds JV-NO might be intended for preventive vaccination of different population groups against viral hepatitis. The use of combination vaccines with reduced antigenic load will reduce the frequency of registration of General and local reactions, and can reduce the cost of vaccination against hepatitis a and B. When this perspective is combined adjuvanted vaccines with reduced antigenic load for persons with reduced levels of immunity, as well as for people living in ecologically unfavorable regions.

The implementation in practice of Depakine with SP-NO will significantly increase the coverage of the vaccination against viral hepatitis compared with the use of plain products, reduce the procedure of vaccination and the cost of protection at the same time against two infections.

Preparation of sample Giacconi against hepatitis a and hepatitis b (A plus).

Depakine can be made on the basis of the substance vacc who are hepatitis a and hepatitis b vaccine by mixing semis followed by the addition of concentrated solution of JV-NO.

For the production of Depakine use: 1) suspension of inactivated hepatitis a virions (strain FOREHEAD-86 or BWA-07), grown in culture transplantable cells 4647, with an activity of 80 ELISA units/ml, and 2) the antigen hepatitis b - HBsAg protein, synthesized by recombinant strain of yeast containing the antigenic determinant of the surface antigen of hepatitis b virus (subtype ayw) in a concentration of 20 µg/ml, adsorbed on aluminium hydroxide; 3) SP-NO (option SP-NO-3-1). To obtain 100 ml of Depakine with SP-NO in aseptic conditions in the dark bowl of the above semi-finished products vaccines hepatitis a and b are mixed in a ratio of 1:1, then with constant stirring portions add 20 mg of SP-NO and left to stir for at least 15 minutes.

To study selected 3 sample Depakine "a+b" (G-1, G-2, G-3) with the content of the component of the hepatitis b virus, 10 μg, a component of the hepatitis a virus 40 - ELISA units and 200 µg SP-NO, different blending technology components.

Example 21. The study of post-vaccination immunity against hepatitis a and hepatitis b

The study of post-vaccination immunity against hepatitis a and hepatitis b are conducted to assess the specificity of the immune response in different ways vaccines against hepatitis a and hepatitis C.

Selections of the candidate vaccine samples GP-1, GP-2, GP-3, having the same if estvo antigens and immunomodulator SP-NO, but different contents of aluminium hydroxide and method of preparation of candidate vaccine preparation. As Comparators use a commercial vaccine containing the approved dosage of viral components approved for medical use:

"hepatitis b vaccine" content viral component 20 µg (DPP-4);

a mixture of antigens of hepatitis b in the amount of 20 µg and antigen of hepatitis a in the amount of 80 ELISA without SP-NO (SE-5).

For immunization using mice of balb/c, male, age 6-8 weeks, weight 18-20 g

Drugs injected subcutaneously at 2 points in a volume of 1 ml From each group of 6 animals subjected to immunization once (with fence serum on day 14), 6 animals twice (with fence serum at 28 day) and the remaining 6 animals of each group is a three by sampling serum on day 42 after the first immunization. For detection of anti HBs antibodies using commercial diagnostician "DS-EIA-ANTI-HBs" (NGO "Diagnostic system").

The results of the evaluation of the immune response, are shown in table 12, show that immunization with different candidate vaccines against hepatitis a and b causes the formation of anti-HBs antibodies in all animals after the first injection of the drug, while the optical density (OD) of all sera exceeded OP to eticheskuyu. Evaluation of immune response to an inactivated hepatitis a virus showed that in the group of candidate vaccines, antibodies were detected in all animals after each injection of vaccine as well as group commercial products.

Antibody titers caused by different variants of candidate vaccines and control preparations containing twice as large doses of viral components (monotherapy HBs antigen with aluminum hydroxide and Depakine containing HBs-antigen, inactivated hepatitis a virus and aluminium hydroxide), comparable.

Example 22. Evaluation of the immunogenicity of the conjugates and complexes SP-NO with hemagglutinin (HA) and neuraminidase (NA) in animals with different genotypes.

Getting vaccinology compounds for example, the conjugates and complexes SP-NO surface protective antigens of influenza virus are carried out by the above-described known methods using the azide method conjugation or complexation.

One of the objectives of the adjuvant is to strengthen the immune response of genetically weakly reactive specimens. In other words, adjuvant, having immunostimulating properties, dramatically enhances the immunogenic properties of the antigen and induces a high immune response in genetically low-reactive individuals to the level of highly reactive. In this work, carried out assessment of the ka opportunities phenotypic correction of immunoadjuvants SP-NO (500 μg) antibody response of mice of different lines on one - and two-immunization

1) the hemagglutinin of influenza virus in a dose of 1 mcg;

2) the hemagglutinin in the same dose in combination with SP-NO;

3) conjugate 1 µg of hemagglutinin with SP-NO. In the experiments used adult mice lines A/Sn, B10CW, CC57W, IAS.

In the graphs of figure 2 presents the results of comparative studies of the production of anti-influenza antibodies after primary and secondary immunization, where the ordinate axis shows the level of anti-influenza antibodies in the serum of mice of different lines in units of optical density of the serum after single and repeated (1-2 on the x-axis) the introduction of the drug:

a) hemagglutinin of influenza virus;

b) hemagglutinin with Polyoxidonium;

C) conjugate hemagglutinin with SP-NO.

When immunization pure hemagglutinin greatest of antibodies registered in mice lines B10CW, the lowest - in mice lines IAS and CC57W after the second immunization. Mouse strain A/Sn occupied an intermediate position. With the introduction of antigen in complex with SP-NO marked enhancement of immune response in mice of all lines in different degrees of severity: the production of antibodies has increased dramatically in A/Sn, expressed - in mice of CBA and to a lesser degree - reacting strongly to the antigen line - B10CW.

In the case of immunization of mice conjugate with SP-NO antibody titer in mice of all investigated lines has increased markedly actually is about the same values (the data presented in table 13).

The data presented in table 13 confirms that the inclusion of immunoadjuvant SP-NO in the vaccine composition of the drug (in the form of a complex or conjugate) leads to the correction of genetically determined humoral immune response: the level of antibody productions in individuals with low reactivity to the antigen increases significantly, while the immune response in highly reactive species increases to a lesser extent compared with the response to "clean" the antigen.

Thus, the copolymers of heterochain hetero-chain aliphatic amines and aliphatic N-oxides (SP-NO) are water-soluble, non-toxic synthetic high-molecular compounds with a high degree of polarity and adsorption ability, able to destructivity on low molecular weight fraction and easily excreted from the body in a wide range of doses activates macrophage link, humoral immune response, increases the resistance of cell membranes to the cytotoxic action of possess a wide spectrum of pharmacological properties, crucial for creation on their basis of drugs and vaccines, high bioavailability, capacity for excretion from the body, by the security application.

Table 2
Pharmacokinetic parameters SP-NO
Pharmacokinetic parametersDesignationUnitsSP-NO 3-1SP-NO 2-1SP-NO 1-1
The maximum concentration in plasmaCmaxMg/ml17,5±0,2021,4±0,619,8±0,3
The time to reach maximum concentration in the blood plasmaTmaxhour0,65±0,10,83±0,150,43±0,1
The period of poluraspredelenia fast α-phaseT1/2άhour0,44±0,060,56±0,070,31±0,05
The period of poluraspredelenia slow β-phaseT1/2βhour36,2±7,7843,2±8,324,6±6,17
The average residence time of drug in the bodyMRThour38,7±6,7557±8,527,1±5,3
Relative bioavailabilityf%89,4±5,7781,6±7,295,4±3,1

Table 3
Detoxification properties SP-NO
GroupDose (mg/kg MRM SP-NO)Put volume, ml/20 gThe number of mice in groupMortality dead/alive
Control--0,560/6
MPM5-0,2563/3
MPM-0,566/0
MPM+SP-NO-3-110100,562/4
MPM+SP-NO-3-210500,561/5
MPM+SP-NO-1-210300,561/5
MPM+SP-NO-2-110200,562/4

Table 4
Antidote properties SP-NO-3-1 when exposed to lethal doses of copper salts
Groups of animalsDose, mg/kg% mortality within 1-2 daysSurvival of jivotina 30 days ( %)
CuSO4SP-NO-3-1
1--0100
212,5-1000
325,0-1000
450,0-1000
512,512,50100
625,025,00100
750,050,00100

Table 5
Detoxification and membranostabiliziruyuschim the properties of the SP-NO (on the model of hemolysis of erythrocytes under the influence of silica)
no groupsThe analyzed connectionHemolysis, %Protection against hemolysis, %
Name of productDose, mg/ml
1The Hanks solution3 ml100,00
2SP-NO-3-2533,3*6b,7*
31019,7*80,3*
4505,4*94,6*
5Gemodez100066,6the 33.4
61500017,9*82,1*
7Poliglyukin1500082,8 17,2
8Albumin10082,018,0
9100013,5*86,5*
* indicated statistically significant differences with group 1 at the lowest value of the significance level of p<0,01

td align="center"> 1,0
Table 6
Immunostimulirutuyu activity of SP-NO
MedicationThe parameters of medicationDose, mg/mouseThe coefficient of stimulationRoute of administration
SP-NO-1-1n=620, q=0,35n, z=0,65n100,06,2subcutaneously (s/C)
SP-NO-1-2n=10, q=0,9n, z=0,1n1000,03,2intraperitoneally (b)
SP-NO-1-3n=800, q=0,2n, z=0,8n6,3n/a
SP-NO-2-1n=650, q=0,2n, z=0,8n10,06,1n/a
SP-NO-2-2n=600, q=0,6n, z=0,4 n500,04,4in a/b
SP-NO-2-3n=850, q=0,5n, z=0.5 n200,05,3n/a
SP-NO-3-1n=700, q=0,25n, z=0,75n50,06,5n/a
SP-NO-3-2n=200, q=0,1n, z=0,9n100,0the 3.8in a/b
SP-NO-3-3n=900, q=0,5n, z=0.5 n20,06,8n/a

Table 7
Characteristics of the obtained variants of TB vaccines
Dose AK, μg/mouseDose of protein, ág/mouseDose of SP-NO, μg/mouseThe ratio AK/SP-NOThe ratio of protein/SP-NO
The drug 11005,05001:51:100
The product 2502.55001:101:200

Table 8
The inoculation of bacilli from the lungs and spleen of infected mice after injection
no groupThe designationThe number of CFU in the lungsThe number of CFU in the spleen
1The drug 1(3,2±0,7)×107(3,2±0,7)×106
2The product 24,2±1,2)×10 7(2,6 ą 0.5)×106
3Drug 3(6,2±0,4)×107(5,2±1,1)×106
4Control(5,6±1,6)×107(8,1±2,6)×106

Table 9
Survival of vaccinated animals
no groupThe number of animals in groupThe designationThe average lifetime (days)
19The drug 144±4,8
27The product 2>64
39Drug 332±5,3
411BCG (Prague) 10653±7,7
5 9Control28±1,6

Table 10
The protective properties of the series of conjugates VI-antigen with SP-NO
MedicationDose immunization mcg VI-PS/mouseThe dose of infection (The No. 4446)The percentage of surviving animals
Sample 151,2100
3,6100
10,8100
Sample 251,2100
3,6100
10,825
VI-PS51,240
3,625
10,8 0
Commercial vaccine51,2100
3,640
10,80

Table No. 11
Thermal response in rabbits after the introduction of commercial vaccines and vaccine VI-PS JV-NO
No. KrolMedicationDose, mcgMeasurement (t°C)
source.1 hour2 hours3 hoursday
1.Sample No. 1100039,439,139,2539,4539,1
2.100038,839,1 39,739,5538,9
3.100038,639,639,4539,5538,6
4.Sample # 220038,8538,638,739,0538,7
5.20039,0539,038, 939,038,6
6.20038,438,838, 95to 38.338,4
7.Vaccine4039,140,5539,739,9538,1
. chemical4039,039,839,7539,239,8
9.Sorbian.4038,240,139,439,2539,1

Table 12
Antibody titers caused by different vaccine candidates
GroupGP-1GP-2GP-3DPP-4GP-5
OPOP/OpcritOPOP/OpcritOPOP/OpcritOPOP/OpcritOPOP/Opcrit
After the first immunization
Cf. value.1,17,80,74,51,07,21,17,70,96,5
Off0,906,10,21,20,8of 5.41,06,70,32,0
After the second immunization
Cf. value.2,416,62,718,92,718,62,416,42,819,1
Off0,64,00,32,22,40,32,20,31,8
After the third immunization
Cf. value.3,020,62,919,82,819,63,020,72,617,7
Off0,00,30,21,20,32,00,00,00,53,4

Table 13
Antibody mice of different lines through 2 weeks after the second immunization preparations containing the hemagglutinin of influenza virus (antibody titer in ELISA)
Line of miceThe average titer of antibodies to the heme is glutinin when immunization of mice
The hemagglutinin (HA)The complex HA with immunoadjuvant SP-NOConjugate HA with immunoadjuvant SP-NO
CC57W57052993342408995
B10CW4891785619182767209
IAS570522621441290948
A/Sn1613697414553913424

1. Copolymers hetero-chain aliphatic poly-N-oxides of the General formula (I):

where R=N, CH;
x=2-4; y=0, 2; n=10-1000; q=(0,1-0,9)n; z=(0,1-0,9)n with pharmacological activity.

2. The copolymer according to claim 1, characterized in that R=CH; x=4, y=0, has the formula (2):
,
is a copolymer conidine and N-oxide conidine.

3. The copolymer according to claim 1, characterized in that R=CH; x=2, y=2, has the formula (3):
,
is a copolymer of hinoklidina and N-oxide of hinoklidina.

4. The copolymer according to claim 1, characterized in that R=N; x=2, y=2, has the formula (4):
,
is a copolymer of triethylenediamine and N-oxide of triethylenediamine.

5. The copolymer according to claim 1, having an antioxidant effect.

6. The copolymer according to claim 1 or 2, or 3, or 4, has a therapeutic effect as a detoxifier.

7. The copolymer according to claim 1, having a therapeutic effect as an immunomodulating agent.

8. The copolymer according to claim 1, which represents immunoadjuvant.

9. The copolymer according to claim 1, which represents an immunogenic carrier antigen or drug.

10. Vaccinifolia means comprising an antigen and an immunogenic carrier, characterized in that it contains as an immunomodulating media copolymer according to claim 1.

11. Vaccinifolia tool of claim 10, characterized in that it is a conjugate of the antigen with a copolymer according to claim 1 in the presence in the structure of the antigen reactive functional groups.

12. Vaccinifolia tool of claim 10, characterized in that it is a compound resulting from the complexation of the antigen with a copolymer according to claim 1.

13. Vaccinifolia tool of claim 10, characterized in that it is a composition obtained by mixing the antigen with a copolymer according to claim 1.

14. The vaccine against hepatitis a and hepatitis b, characterized in that it contains in czisny drug containing both AG CAA and HBsAg or vaccine preparations against hepatitis a and hepatitis b and copolymer hetero-chain aliphatic poly-N-oxides according to claim 1.

15. The vaccine according to 14, characterized in that it contains as AG HAV antigens derived from strain FOREHEAD-86 hepatitis a virus in the culture of transplantable cells 4647, and the content of components in the dose is:

AH HAV40-60 ELISA units
HbsAgthe 2.5-20 mcg
SP-NO0,1-10 mg

16. Vaccine composition comprising the vaccine formulation and immunoadjuvant, characterized in that it contains as immunoadjuvant copolymer according to claim 1.

17. The drug, including drug substance and a carrier, characterized in that it contains as a carrier of the copolymer according to claim 1.

18. Drug at 17, characterized in that it is a conjugate of medicinal substance with a copolymer according to claim 1 in the presence in the structure of the drug substance reactive functional groups.

19. Drug at 17, characterized in that it is a compound obtained by reaction of compleksoobrazutee the drug substance with a copolymer according to claim 1.

20. Drug at 17, characterized in that it is a pharmaceutical composition obtained by mixing a medicinal substance with a copolymer according to claim 1.



 

Same patents:

Chemical process // 2399633

FIELD: chemistry.

SUBSTANCE: method involves reaction of a nitrogen-containing precursor selected from a group consisting of amines, polyamines, polyaminoamides, polyurethanes and mixtures of said substances with epihalohydrin. The method also involves a step for cleaning the formed polymer solution from organic impurities. The cleaning step involves separation of impurities from the epihalohydrin polymer solution through extraction of said impurities. Extraction is carried out using CO2 in liquid, supercritical or close to supercritical state. The extracted impurities are then separated.

EFFECT: efficient removal of organic impurities from the obtained reaction product.

14 cl, 5 ex, 2 tbl, 2 dwg

FIELD: chemistry.

SUBSTANCE: invention relates to a method of preparing an aqueous polymer dispersion. The method involves reacting one or more cationic or non-ionic monomers (m) and a polymer (B) in the presence of a co-stabiliser A. Polymer (B) is hydrolysed in the presence of a co-stabiliser (A) with formation of a polymer stabiliser (C). The obtained aqueous dispersion is further used as a holding agent in paper production.

EFFECT: disclosed dispersion demonstrates good retention and dehydration properties.

12 cl, 6 ex, 1 tbl

FIELD: chemistry.

SUBSTANCE: invention relates to use of polymers as a wetting agent in offset printing. The wetting agent contains at least one polymer P, which is a product of reacting polyethyleneimine with a cross-linking agent obtained by reacting polyethylene glycol with molecular weight 1500 with epichlorohydrin, and acrylic acid and contains amino groups modified with a -CH2CH2COOH group and/or its salts. Described also is an offset printing method which uses the said wetting agent.

EFFECT: wetting agent ensures formation on non-printing parts of the printing plate of a stable water film which prevents movement of printing ink to the said parts during printing.

9 cl, 2 tbl, 1 ex

FIELD: chemistry.

SUBSTANCE: invention relates to use of cationic polymers, specifically to a method of neutralising clay in sand. The method neutralising clay in sand, designed to obtain hydraulic compositions, comprises steps for adding a cationic polymer into the compositon or into one of its components. The said cationic polymer has cation charge density of over 0.5 meV/g, and characteristic viscosity less than 1 dl/g.

EFFECT: use in accordance with the invention enables efficient neutralisation of clay in hydraulic compositions.

24 cl, 4 tbl, 24 ex

FIELD: oil and gas production.

SUBSTANCE: method of modifying water permeability of underground formation consists in pumping water composition into underground formation. Water composition contains 0.005-2 wt % of poly-hydroxietheramine, replaced with alkyleleneoxide or its salt, where the said poly-hydroxietheramine is produced by interaction of diepoxide out of group containing of diglycidile ethers of polyatomic phenol with one or several amines containing two reaction-capable atoms of hydrogen and not necessary by interaction of thus produced polyhydroxietheramine with acid or alkylation agent of formula R14X, where: R14 - C1-C4 alkyl and X are halogen, sulphate or sulphanol with formation of salt. Water composition contains 0.005-2 wt % of the above said water soluble polyhydroxietheramine or its salt and 0.005-2 wt % of listed solvents. The invention is developed in dependant claims of formula.

EFFECT: reduced contents of water in products, raised efficiency of extraction of oil out of formation.

25 cl, 4 tbl, 2 ex

FIELD: chemistry.

SUBSTANCE: invention relates to the heteropolymer salts of hexamethyleneguadinine and method for obtaining it, it is used as a means of disinfecting in medicine, veterinary science, for disinfection of natural and waste water, for protection of the materials of plant and animal origin, for example, of wood, cotton the skin, fur from biodegeneration, as well as in other sectors of national economy, where biocidal products are required. Heteropolymer salts of hydrochloride, phosphate and sodium dihydroorthophosphate of hexamethyleneguadinine of the following formula: , where: A=5-25 mol%; B=60-90 mol%; C=5-30 mol%, can be obtained in two ways. The first method of obtaining heteropolymer salts consists in adding and mixing up of the base of poly- hexamethyleneguadinine, dissolved in ethyl alcohol or in water, with hydrochloric acid, phosphoric acid and 30% aqueous solution of sodium dihydroorthophosphate, or with hydrochloric acid, phosphoric acid and aqueous sodium hydroxide. Then the obtained heteropolymer is separated and fried. Initial components are take in the following molar relationship base poly-hexamethyleneguadinine: hydrochloric acid: phosphoric acid: solution of sodium dihydroorthophosphate - 1: 0.05-0.25 : 0.60-0.90 : 0.05-0.30, or in the following molar relationship of the base poly- hexamethyleneguadinine: hydrochloric acid; phosphoric acid; solution of sodium dihydroorthophosphate - 1: 0.05-0.25 : 0.65-1.20 : 0.05-0.30. The second method of obtaining the heteropolymer salts consists in that to the hydrochloride of poly- hexamethyleneguadinine, dissolved in ethyl alcohol, alcoholic solution of sodium hydroxide obtaining the mother liquor, to which phosphoric acid and 30% aqueous solution of sodium dihydroorthophosphate is added while mixing. The obtained heteropolymer is dried. The initial components are put in the following molar relationship: hydrochloride of the poly- hexamethyleneguadinine; alcoholic solution of sodium hydroxide: phosphoric acid: solution of sodium dihydroorthophosphate - 1: 0.75-0.95 : 0.60-0.90 : 0.05-0.30.

EFFECT: invention enable to obtain heteropolymer salts with high antimicrobial activity.

4 cl, tbl, 7 ex

FIELD: inorganic chemistry.

SUBSTANCE: invention covers water soluble sizing compounds providing oil resistance/grease- and waterproofness applied for cellulose materials processing, containing repeated elements including polyamine with at least 3 nitrogen atoms in every repeated monomer element where amides are partially or completely substituted with a. cellulose interacting group resulted from reaction of polyamine and at least one reactive chemical compound selected from number including epihalohydrin, epiorganosulfonates and oxirane implying that interaction with cellulose arises after polyamine reaction or due to further reactivation; b. hydrophilic and hydrophobic fluorine-containing chemical group substituted with alkyl chain containing hydrophilic group; besides cellulose interacting group cross-links polyamine groups, and substance molecular weight Mw is from 10000 to 35000000 daltons determined by gel-penetrating chromatography relative to, polymethylmethacrylate standard. In addition invention covers chemical substance containing polymer composition with product of interaction between fluorocarbon-substituted polyamine of IV structure or epihalohydrin or epiorganosulfonate; method of sized paper production providing oil resistance/grease- and waterproofness (versions); paper production (versions); aqueous dispersion (versions). This substance effectively works providing oil resistant/grease- and waterproof sizing at high temperatures.

EFFECT: sizing compound production providing oil resistance/grease- and waterproofness of cellulose materials.

36 cl, 20 tbl, 125 ex

FIELD: chemical technology.

SUBSTANCE: invention relates to a method for synthesis of conducting polyaniline without adding any acid to the reaction. Method involves dissolving oxidizing agent in pure solvent and addition of aniline monomer at stirring by drops. Then the reaction mixture is kept at temperature 10-35°C for 4-10 h. Prepared conducting polyaniline is precipitated by immersion in distilled water and separated. Invention provides development of noncorrosion method for synthesis of polyaniline that shows safety for environment.

EFFECT: improved method of synthesis.

12 cl, 4 ex

FIELD: organic chemistry, chemical technology.

SUBSTANCE: invention describes a method for synthesis of organic-soluble polyguanidine salt. Method involves preparing polyguanidine base by the alkaline dehydrochlorination reaction of polyguanidine hydrophilic salt followed by neutralization of polyguanidine base with hydrophobic organic acid. As hydrophobic organic acid pelargonic, benzoic or undecylenic acid are used, and polyhexamethylene guanidine hydrochloride or poly-(4.9-dioxadodecane guanidine) hydrochloride are used as polyguanidine hydrophilic salt. Also, invention describes polyguanidine organic-soluble salt possessing biocide and hydrophobic properties of the following structural formula: wherein n = 5-70; R means ˜(CH2)6˜, ˜(CH2)3O(CH2)4O(CH2)3˜; R1 means CH3(CH2)4CH=CH(CH2)4˜, ˜CH3(CH2)7˜.

EFFECT: improved method of synthesis.

3 cl, 2 tbl, 4 ex

FIELD: organic chemistry, polymers, chemical technology, biotechnology.

SUBSTANCE: method involves carrying out the oxidative polymerization reaction of aniline for a single step. Laccase that uses air molecular oxygen as a substrate is used a s catalyst for reaction wherein in process of this reaction oxygen is reduced to water. After carrying out the enzymatic polymerization reaction a formed polyaniline precipitate is separated from reaction solution, washed out with water and, if necessary, the dedoping process of emeraldin conducting polyaniline salt with ammonia aqueous solution is carried out. Advantage of method involves it's a single step in carrying out the process and ecological purity of the end product. Invention can be used in creature of bio- and chemosensors and separation of optically active compounds.

EFFECT: improved method of synthesis.

5 ex

FIELD: chemistry.

SUBSTANCE: invention describes a method for industrial production of modified polymers in accordance with which a ground polymer undergoes gas-chemical modification in fluidised state using reactive gases, including inert gases and ozone as a gaseous oxidising agent and the obtained activated powder is mixed with a monomer or a group of monomers or a polymer, followed by their copolymerisation through surface-initiated polymerisation in molten stage in an extruder. The method is characterised by that gas-chemical modification is carried out at temperature not higher than 30°C and ozone concentration of 5-15 wt %, while blowing with reactive gases for 10-30 minutes, and polymerisation in the extruder is carried out at 130-230°C, while augers are rotating at 300-500 rpm, L/D of not less than 40, for synthesis of graft and block copolymers with degree of grafting of not less than 80%. The invention also describes a device for realising said method.

EFFECT: cost-effective and environmentally clean industrial method of modifying the surface of polymers with reactive gases with subsequent synthesis of graft and block copolymers through surface-initiated polymerisation.

5 cl, 4 dwg, 5 tbl, 7 ex

FIELD: chemistry.

SUBSTANCE: invention refers to method for making unsaturated polyketones of individually variable molecular weight and carbonyl group content. Disclosed method for making unsaturated polyketones is ensured by isoprene copolymers oxygenation with nitrous oxide (N2O) with diene and/or vinyl monomers at 150-300°C and nitrous oxide pressure 10 to 100 atm. Besides, oxygenation can be enabled with mixed N2O and solvent gases, such as inert gases, alkans C1-C4 or their mixtures. Offered method results in controlled macromolecule C=C bond breakage rupture in initial polymer.

EFFECT: making unsaturated polyketones of specified molecular weight and carbonyl group content.

9 cl, 18 ex

FIELD: chemical industry; the methods and installations for production of the oxidized atactic polypropylene.

SUBSTANCE: the invention is pertaining to production of the oxidized atactic polypropylene with the molecular mass of 5500-38000 and the polar functional groups, which may be used as the component of the various composite materials, the multipurpose additives for lubricants, anticorrosion coatings. For production the above-indicated oxidized atactic polypropylene they use the industrial atactic polypropylene with the molecular mass of 20000-40000 and the oxidization conduct at least in two stages with the adjustable temperature drop from 250 to 150°С within 1-6 hours at the air consumption of 0.6-1.9 l/(minute·kg). The installation for the method realization contains at least two in series connected reactors supplied with the electric heater at the ratio of the height of the reactor to its diameter laying within the limits of 2.0-5.0, the device for the air supply is made in the form of the vertical pipe with the air supply through its upper part and in the lower part it is supplied with the nozzle with the slits arranged along the perimeter of the lower edge. At that the diameter of the nozzle with respect to the diameter of the reactor is chosen within the limits of 0.25-0.5.

EFFECT: the invention ensures production of the oxidized atactic polypropylene with the molecular mass of 5500-38000 and the polar functional groups, which may be used as the component of the various composite materials, the multipurpose additives for lubricants and the anticorrosion coatings.

5 cl, 1 dwg, 4 ex

FIELD: polymer production.

SUBSTANCE: invention provides polymers containing functional carbonyl groups, which are prepared via oxygenation of double bonds C=C of polymers with the aid of nitrogen monoxide in presence of diluent gas: individual C1-C4-alkane or C1-C4-alkane mixture, followed by conversion of carbonyl groups into another-type groups.

EFFECT: increased productivity of process while ensuring compete blastproofness.

4 cl, 1 tbl, 9 ex

FIELD: chemical industry; methods of production of the polymeric compounds and the oligomers containing the functional groups.

SUBSTANCE: the invention is pertaining to the method of production of the polymeric compounds or the oligomers containing in their compositions the carbonyl functional groups. The invention presents the method of production of the polymeric compounds or the oligomers containing the functional carbonyl groups by oxygenation of the double bonds ofC=C polymeric and oligomers by means of nitrous oxide at presence in the capacity of the gas-diluent of one of the alkanesC1-C4 or their mixture with the subsequent possible transformation of the carbonyl groups into the functional groups of other type. The process may be conducted at presence of the stabilizing agents increasing the thermal stability of the polymeric compounds or oligomers. The technical result of the invention is the increased efficiency of the process at provision of the complete explosion safety.

EFFECT: the invention ensures the increased efficiency of the process at provision of the complete explosion safety.

6 cl, 1 tbl, 16 ex

The invention relates to a method for modification of polymers by introducing them in oxygen-containing functional groups

The invention relates to the modification of polymers containing carbon-carbon connection

FIELD: medicine.

SUBSTANCE: invention refers to medicine, and can be used if extremity vein catheterisation is required for multiple intravenous introduction of drug solutions. That is ensured by vein catheterisation followed by change of an extremity position with respect to a patient's trunk so that an open pavilion of the catheter is arranged below a level of a patient's right atrium. An infrared imager is used to monitor an extremity surface condition within a projection of the catheter. If observing a region of local hyperthermia along the vein, its content is examined by ultrasound. A thrombus if any is sized up; tourniquets are applied below and above the region of local hyperthermia to complete sanguimotion cessation in the vein. Then, blood is taken from the vein to its devastation, and the volume of taken blood portion is measured that is followed by introduction of 2% lidocaine hydrochloride 0.5 ml and a thrombolytic solution at temperature +42°C in the volume necessary for complete vein filling. 3 minutes later, the whole liquid vein content is removed through the catheter, and the tourniquets are released, the vein content is examined by ultrasound. The presence of a thrombus requires another 3-minute tourniquet application with venous blood replacement by a local anaesthetic solution with a thrombolytic agent to stabilise thrombus size, then the catheter is removed, and the drugs are introduced by means of another catheter which is inserted in another extremity vein.

EFFECT: method allows higher safety and effectiveness of vein catheterisation due to early phlebitis detection and exact delivery of fibrinolytic agents to the thrombus to dissolve a "fresh" portion of the thrombus that allows reducing a risk of thromboembolism.

1 ex

FIELD: medicine.

SUBSTANCE: what is offered is administration of haemorrheologic agents presented by water-soluble hybrid macromolecular phenol antioxidants O-(3-(3,5-di-tert-butyl-4-hydroxyphenyl)propenyl)-dextrane, O-(3-(3,5-di-tert-butyl-4-hydroxyphenyl)propenyl)-O-(2-hydroxyethyl)-starch, polyethylene glycol bis-3-(3,5-di-tert-butyl-4-hydroxyphenyl)-propionate, O-[2-benzamido-3-(3,5-di-tert-butyl-4-hydroxyphenyl)acryloyl)]-dextrane, O-[2-benzamido-3-(3,5-di-tert-butyl-4-hydroxyphenyl)acryloyl)]-O-(2-hydroxyethyl)-starch prepared with using hydroxyl-containing dextrane polymers of molecular weight 40 kDa or hydroxyethylised starch of molecular weight 200 kDa or polyethylene glycol of molecular weight 20 kDa. The blood viscosity increase limited by the listed substances is shown.

EFFECT: improved properties of the agent.

1 tbl, 12 ex

FIELD: chemistry.

SUBSTANCE: invention relates to a method of producing prasugrel hydrochloride of the formula:

,

with low content of "ОХТР", involving preparation of free prasugrel containing "ОХТР" from 2-silyloxy-5-(α-cyclopropylcarbonyl-2-fluorobenzyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine, dissolving the obtained free prasugrel in an inert solvent and optionally adding hydrochloric acid in drops to the solution for reaction.

EFFECT: novel method of producing prasugrel with low content of impurities, specifically "ОХТР" by-product.

2 cl, 6 dwg, 1 tbl, 2 ex

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