Method for preparing stabiliser-coated nanocrystalline cerium dioxide

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to a method for preparing stabiliser-coated nanocrystalline cerium dioxide characterised by antioxidant activity. The method involves preparing an aqueous solution of cerium salt and a stabiliser representing maltodextrin with a molar ratio of cerium to the stabiliser 1 to 1-4. Then, the prepared aqueous solution is added with drops of hydrous ammonia with stirring, and pH of the prepared solution is gradually increased to 7-8, maintained for 1-4 hours, the prepared colloidal solution of hydrous cerium nanoparticles is added with hydrous ammonia, and pH is increased to 11-12 and maintained for 1-10 hours to form a colloidal solution of cerium dioxide. Thereafter, an alcohol or ketone excess is removed and brought to the boiling point, while the formed precipitate of the non-aggregated nanoparticles of stabiliser-coated cerium dioxide, separated by decantation or filtration, washed 1-4 times in alcohol or ketone, and dried at temperature 50-80°C to constant weight. The prepared powder of the non-aggregated nanoparticles of stabiliser-coated cerium dioxide is re-dispersed in a polar solvent to form aggregation resistant sol.

EFFECT: invention provides preparing stabilised nanocrystalline cerium dioxide with a hydrodynamic diameter of 6-10 nm.

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The invention relates to chemical technology, nanotechnology and biotechnology, namely antioxidant drug on the basis of nanocrystalline cerium dioxide and stabilizer, designed to protect the cells of a living organism from oxidative stress, in particular to the method of its production.

Oxidative stress is involved in the pathogenesis of more than 100 different diseases - primarily neurodegeneration, such as Alzheimer's disease and other types of dementia, Parkinson's disease, amyotrophic lateral sclerosis, epilepsy and multiple sclerosis. In addition to these diseases, oxidative stress is considered to be a participant other disorders - arthritis, cardiac disorders and diseases that cause irreversible blindness, such as diabetic retinopathy, macular and retinal degeneration. Finally, the aging of any organism is accompanied by oxidative stress, because the activity of the natural antioxidant system is reduced with age, and the concentration of products of lipid peroxidation is increased. Thus, due to the disruption of the antioxidant system of the body or a significant increase in the level of exogenous reactive oxygen species possible very severe consequences. In such situations to protect the body need different kinds of Antioch identy.

On the other hand, reactive oxygen species are involved in cell metabolism, particularly in the respiratory and energy processes, as well as components of protective mechanism against foreign factors, including viruses and bacteria. Thus, the challenge is targeted regulation of reactive oxygen species in the cell.

It is known that nanocrystalline ceria exhibits high antioxidant activity [Vchebanov, Aberbach and Averatec, USP 78, (9), 924 (2009)], however, in the work there are no instructions on how to obtain pharmaceutically acceptable dosage forms stable containment of cerium dioxide.

Known also plenty of ways to get covered with a stabilizing shell of nanocrystalline cerium dioxide, but only few of them meet the criteria to pharmaceutically acceptable dosage forms. Characteristics of the drugs used are determined not only by the particle size of nanocrystalline cerium dioxide and the condition of its surface, but also by the properties of the stabilizing shell.

Thus, the known method of synthesis of water-soluble biocompatible stable Zola nanocrystalline cerium dioxide [A.S.Karakoti, N.A.Monteiro-Riviere, R.Aggarwal, et al., J. Minerals, Metals and Materials Soc. 6 (3), 33 (2008)]. The disadvantage of this method is that it offers no practical method to set the size of the particles of cerium dioxide for regulation of antioxidant activity of the final product in relation to the active forms of oxygen.

In [Y..Tsai, J.Oca-Cossio, K.Agering, et al., Nanomed. 2 (3), 325 (2007)] as a stabilizer of nanocrystalline cerium dioxide used non-toxic lecithin. The particle size of CeO2in the resulting ash was 3.5 nm, while the Sol was stable in buffer solution triacrylate. To the disadvantage of the proposed method include the fact that the use of lecithin as a stabilizer prevents redispersible anhydrous product to obtain a stable water Zola.

In addition, no proposed practical method to set the size of the particles of cerium dioxide for regulation of antioxidant activity of the final product in relation to the active forms of oxygen.

In [S.Maensiri, .Masingboon, P.Laokul, et al., Crystal Growth & Design, 7 (5), 950 (2007)] as a stabilizer of nanocrystalline cerium dioxide was used albumin chicken egg. Unfortunately, the resulting product was unstable and subjected to aging due to degradation of the stabilizer, so it is not of practical interest.

In [N.Izu, I.Matsubara, .Itoh, et al., Bull. Chem. Soc. Japan 81 (6), 761 (2008) stabilization of nanocrystalline cerium dioxide used non-toxic polyvinylpyrrolidone (molecular weight 4250÷18000 g/mol), however, the use of such stabilizer led to the fact that the size of the obtained particles CeO2turned out to be too large (50÷100 nm), and therefore the resulting Sol of nanocrystalline cerium dioxide showed a lack of antioxidant activity.

Known invention [WO/2008/030815], in which the stabilizer of nanocrystalline cerium dioxide using 2-[2-(2-methoxyethoxy)ethoxy]acetic acid or ethylenediaminetetraacetic acid.

The use of the proposed stabilizers prevents redispersible anhydrous product to obtain a stable water Zola, which limits its practical use. In addition, the use of this stabilizer is not possible to adjust the particle size of cerium dioxide and thereby adjust its antioxidant activity compared to the active forms of oxygen.

Known invention [WO/2008/083401], which describes a method of synthesis of nanocrystalline cerium dioxide in reverse micelles using as a stabilizer, bis-2-ethylhexylphthalate sodium (AOT) and the additional stabilization of the obtained particles of sodium citrate.

The proposed method allows the authors to obtain nanocrystalline cerium dioxide with a particle size of from 2 nm to 10 nm. However, the synthesis is carried out in an environment Tox is cnyh organic solvents (hexane, benzene, toluene), which can contaminate the final product and prevent its use for protection of biological objects from oxidative stress. The use of AOT does not allow to dry, long-term storage and redispersible particles of cerium dioxide.

In [A.S.Karakoti, S. V. N. T. Kuchibhatla, K.S.Babu, and S.Seal, J.Phys. Chem. With 111 (46), 17232 (2007)] it is shown, that is covered with a stabilizing shell nanocrystalline ceria can be obtained using as a stabilizer polyhydroxylated connections.

In the invention [US 2010098768] the same authors propose to use as the stabilizer of nanocrystalline cerium dioxide polyethylene oxide, dextran, polyethylenimine, polyacrylic acid, chitosan, alginate, or a combination of both.

Both of the above works do not reveal ways of varying the size of the CERIC oxide particles for targeted regulation of the degree of protection from oxidative stress and does not describe methods of purification of sols. In these works also not received redispersible form nanoparticles stabilized cerium dioxide, which, in our opinion, is connected with agregirovannosti obtained nanoparticles.

The known method [J.M.Perez, A.Asati, S.Nath, and A.Kaittanis, Small, 4, 552 (2008)] get covered with a stabilizing shell of nanocrystalline cerium dioxide used is eaten as a stabilizer biologically valid dextran T10, selected as a prototype.

This method of obtaining includes the preparation of a solution, prepared by mixing 5 ml of a solution of cerium nitrate(III) (Aldrich, 99%) with a concentration of 1.0 M and 10 ml solution of dextran T10 with a molecular weight of 10000 g/mol (Amersham Bioscience), which is a low molecular weight polyhydroxyethyl connection, with a concentration of 1.0 M, the addition of the prepared solution to 30 ml of aqueous ammonia solution (Sigma Aldrich, 30%) and stirring the mixture for 24 h at 25°C, centrifuging the mixture for 30 min at a rotor speed 4000 rpm for separation of coarse aggregate particles, the concentration and purification of the obtained colloidal solution of free dextran T10 filtration Amicon cell (YM 30 K; Millipore Inc.), for subsequent storage at 4°C. the Obtained composition for protection of biological objects from oxidative stress contained nanocrystalline ceria size of 4 nm, covered with a stabilizing shell molecules of dextran T10.

The disadvantage of the prototype is that the high speed of the process of formation of nanocrystalline cerium dioxide, associated with the addition of the cerium salt solution in the ammonia solution, and not Vice versa, it is not possible to adjust the particle size of nanocrystalline cerium dioxide for directional control CTE is the new defense against oxidative stress, and there is no evidence of receipt of the product in the form of a dry powder, which could be completely redispersible in water or another polar solvent, which is associated with agregirovannosti obtained in a colloidal solution of nanoparticles.

The invention is aimed at finding ways to obtain the drug for protection of biological objects from oxidative stress, contains covered with a stabilizing shell nanocrystalline ceria with hydrodynamic diameter 6÷10 nm and the particle size of the actual nanocrystalline cerium dioxide 2÷5 nm, characterized by the possibility of obtaining a product with high antioxidant activity, enabling directed to set the particle size of nanocrystalline cerium dioxide by varying the molar relationship cerium salt and a stabilizer in an aqueous solution for directional control of the degree of protection from oxidative stress, the presence of a simple stage purification from impurities, the stability of the product when stored in a dry powdered state, full redispersible the dried powder in water or other suitable polar solvent, for example dimethyl sulfoxide, formamide, ethylene glycol, glycerin, with the formation of a stable Sol with both low and high concentration accounts for the creation of conditions to prevent aggregation of the particles in a colloidal solution, as well as the simplicity of the process execution.

The technical result is achieved by the method for obtaining covered with a stabilizing shell of nanocrystalline cerium dioxide, characterized by antioxidant activity, which consists in the fact that preparing an aqueous solution of cerium salt and a stabilizer with a molar ratio of cerium and stabilizer 1:1÷4, in terms of the number of monomers in the structure of the stabilizer, the stabilizer is used maltodextrin, to the resulting aqueous solution was dropwise added with stirring an aqueous solution of ammonia and slowly increase the pH of the resulting solution to a value of 7÷8, incubated for 1÷4 hours, to the resulting colloidal solution of nanoparticles of cerium hydroxocobalamine add an aqueous solution of ammonia and increase the pH to 11÷12, and then incubated for 1÷10 hours prior to the formation of a colloidal solution of cerium dioxide, then add an excess of alcohol, selected from a number of: methanol, ethanol, isopropanol, propanol; or ketone selected from the range: dimethylketone, methyl ethyl ketone, diethylketone; and bring to a boil, the precipitate non-aggregated nanoparticles of cerium dioxide coated with a stabilizing shell is separated by decantation or by filtration, washed with 1÷4 times specified alcohol or ketone and dried p and a temperature of 50÷80°C to constant weight, thus redispersion resulting powder non-aggregated nanoparticles of cerium dioxide coated with a stabilizing shell in a polar solvent leads to the formation of aggregately-resistant Zola.

It is advisable that, as a salt of cerium use water-soluble cerium salt with a solubility of not less than 6×10-3mol of cerium in 1 l of water.

It is possible that as the polar solvent used water, dimethylsulfoxide, formamide, ethylene glycol, glycerin.

The essence of the invention lies in the fact that to get a non-aggregated form covered with a stabilizing shell of nanocrystalline cerium dioxide perform a slow increase in pH of the solution containing a salt of cerium and a stabilizer to form hydroxocobalamine cerium, by addition of aqueous ammonia to the solution, and not Vice versa, to create conditions to prevent aggregation of the particles in a colloidal solution, further carry out the curing of the obtained colloidal solution of particles hydroxocobalamine cerium, and then raise the pH of the solution for the conversion of hydroxocobalamine in nanocrystalline cerium dioxide, cerium (NDC). Unlike the prototype clearing covered with a stabilizing shell of nanocrystalline cerium dioxide is conducted directly after the formation of colloidal RA is down, and after the deposition of the obtained aggregated particles by boiling in alcohol or ketone.

The specified technical task and the specified technical result is achieved by using as the stabilizer of a non-toxic, readily available and cheap maltodextrin, forming in all concentrations inviscid solutions.

Maltodextrin is a product of partial acid or enzymatic hydrolysis of potato or corn starch.

It is known that maltodextrin is used as inert excipients in the manufacture of food additives, because according to the FDA it is a safe food ingredient and approved for use in the food industry.

Maltodextrin is a mixture of carbohydrates, having in its chain 2÷20 structural elements of glucose connected by α-(1→4) glycosidic bonds.

Moreover, the stabilizer provides obtaining aggregated particles and stabilization Zola due to the steric barrier to aggregation of the particles due to the spatial separation as nanoparticles hydroxocobalamine cerium during the synthesis and nanocrystalline cerium dioxide.

The size of the resulting particles of nanocrystalline dioxi is and cerium regulate the ratio of the concentrations of the parent compounds of cerium and stabilizer. Sol obtained is covered with a stabilizing shell of nanocrystalline cerium dioxide precipitated and washed repeatedly lower aliphatic alcohols or ketones, dried and stored in appropriate conditions. The purpose of biomedical applications powdery product redispersion in water to form solutions of nanocrystalline cerium dioxide concentration required or used in the form of powder, for example, for oral administration.

The essence of the invention is illustrated by the following accompanying illustrations:

Figure 1. Electron micrograph and the results of dynamic light scattering of colloidal solution obtained according to Examples 1-4.

Figure 2. UV absorption spectra of the obtained colloidal solution according to Examples 1-4.

Figure 3. The width of the forbidden zone is covered with a stabilizing shell of nanocrystalline cerium dioxide obtained according to Examples 1-4.

Figure 4. The change in optical density of the solutions of the dye methyl violet and ferrous iron containing 200 mm covered with a stabilizing shell of nanocrystalline cerium dioxide obtained according to Examples 1-4, with the introduction of hydrogen peroxide. Curve "0" control without entering coated with a stabilizing shell of nanocrystalline cerium dioxide.

Figure 5. Relative if estvo surviving fibroblasts (%), exposed to hydrogen peroxide after processing is covered with a stabilizing shell nanocrystalline cerium dioxide. Non columns correspond to the numbers of Examples 1-4. Column "0" corresponds to a control experiment in which cells were exposed to the effect of hydrogen peroxide in the presence only of the stabilizer maltodextrine without nanocrystalline cerium dioxide.

6. A computer model of particle NDC stable molecule maltodextrin (n=12) with different orientation of the adsorbate. A - vertical orientation, B - planar orientation.

7. The change of absorption spectra of a solution containing the dye methyl violet and Sol ferrous iron, with the introduction of hydrogen peroxide: a - source solution, b is 0.01 ml of N2O2in - 0,02 ml of H2O2Mr. 0.05 ml H2O2(d) 0.1 ml of H2O2.

The present invention is implemented as follows. In the vessel of appropriate volume to prepare an aqueous solution of cerium salt and maltodextrin with a dextrose equivalent DE of 5-20 with a molar ratio of cerium and stabilizer 1:1÷4, in terms of the number of monomers in the structure of the stabilizer. When you use fewer of the stabilizer formed as a result of the synthesis of the Sol is aggregately unstable, use more to what icesta stabilizer is not economically feasible. With vigorous stirring, the pH is slowly increased by using an aqueous solution of ammonia to a value of 7÷8 and is maintained at this level for 1÷4 hours for hydrolysis of salts of cerium (III) and partial oxidation of cerium (III) to cerium (IV). With less duration of this stage, the resulting product when redispersion in water has no aggregative stability. The increase in the duration of this stage is not economically feasible. Reducing the pH of the solution will lead to a significant increase in the duration of the synthesis that is not economically feasible. Increasing the pH of the solution will result in a product containing polydisperse particles of cerium dioxide, which will lead to a significant decrease in antioxidant activity of the final product, as well as to the aggregation of nanoparticles. After that, the pH of the solution increased to 11÷12 and incubated for 1÷10 hours for the formation of nanocrystalline cerium dioxide. With less duration of this stage, the particles of cerium dioxide will not have time to fully secretariats that will lead to a significant decrease in the antioxidant activity of the final product. The increase in the duration of this stage does not change the properties of the final product. Then in the resulting colloidal solution of nanocrystalline cerium dioxide add OFL the current for example, two-volume, lower aliphatic alcohol or ketone and bring mixture to a boil on a water bath. This forms a thick sludge containing particles coated with a stabilizer of nanocrystalline cerium dioxide. The precipitate was separated by decantation or filtration, cleaned from impurities and residues of the original substance by washing 2÷3 times with alcohol or ketone and dried in a drying Cabinet at a temperature of 50÷80°C to constant weight. The resulting powder was stored in a cool, dry place until use. The drug can be used in the form of the following pharmaceutically acceptable dosage form: dry powder; Zola after redispersible dry powder in a polar solvent.

Below are examples of implementations of the claimed invention. The examples illustrate but do not limit the proposed method.

Example 1

1. In a three-neck flask, containing an addition funnel, the electrode of the pH meter and stirrer, was placed 3.7 g (0.01 M) chloride heptahydrate cerium (III), 7.3 g of maltodextrin DE 10-12 (~0.04 M in terms of glucose) and 200 ml of bidistilled water.

2. Turned on the mixer and mixed system until complete dissolution of the components.

3. In an addition funnel volume of 50 ml was placed 1 N ammonia solution, included pH meter and with vigorous stirring to the reaction mixture dropwise EXT the wheelie ammonia solution.

4. Maintained a constant value of pH (pH 8). Transparent colloidal solution thus acquired a straw-yellow color, gradually turning dark brown. The next portion of the ammonia was added after the pH of the solution fell below pH 7.5.

5. After pH of the solution has ceased to decline, the addition of ammonia is stopped and the mixture was stirred for 2 hours.

6. In the reaction mixture is added an aqueous solution of ammonia to a value of pH pH 12 and mixed system within 8 hours.

7. Deleted the addition funnel and the electrode of the pH meter.

8. Introduced in the system 400 ml of isopropyl alcohol.

9. When the stirrer brought the mixture to a boil on a water bath.

10. Stopped stirring and given the opportunity to form a white cheesy draught.

11. Decanted the supernatant and additionally introduced into the flask 100 ml of isopropyl alcohol. Twice repeated procedures 9÷10.

12. The residue was quantitatively transferred into a Petri dish and dried in a drying Cabinet at 60°C until constant weight (about 2÷3 hours).

13. The product yield was about 8 g (~88% of theoretical).

14. The obtained product was stored in a tightly closed bottle in a cool place, protected from direct sunlight.

15. The product is completely redispersible in p is lannom solvent and received medication in the form of aggregately-resistant Zola.

The particle size of nanocrystalline cerium dioxide was 2 nm, and the hydrodynamic diameter of the particles is covered with a stabilizing shell of nanocrystalline cerium dioxide was 6 nm (Figure 1). Absorption spectrum obtained Zola in units of molar extinction are shown in figure 2 (curve 1). The magnitude of the band gap obtained nanocrystalline cerium dioxide amounted to 3.30 eV (Figure 3). The dependence of the optical density of a solution of methyl violet and ferrous iron in the presence of the thus obtained coated nanocrystalline cerium dioxide from the amount added to the hydrogen peroxide solution is shown in Figure 4 (curve 1). The achievement of the technical result, namely antioxidant activity of the obtained preparation is shown in Figure 5.

Additional properties of the dry product (particle size of nanocrystalline cerium dioxide and the width of the forbidden zone), properties of the drug after redispersion in water (hydrodynamic diameter of the particles is covered with a stabilizing shell of nanocrystalline cerium dioxide, protection from oxidative stress) were re-determined after storage for 6 months at 4°C. it was Found that characteristics of the drug after storage did not differ from the characteristics of the original teacher who rata more than 2%.

Example 2

The method of obtaining implemented similar to that described in Example 1, except that the amount of maltodextrin was 5.7 g (~0.03 M in terms of glucose).

The particle size of nanocrystalline cerium dioxide was 3 nm, and the hydrodynamic diameter of the particles is covered with a stabilizing shell of nanocrystalline cerium dioxide was 7 nm (Figure 1). Absorption spectrum obtained Zola in units of molar extinction are shown in figure 2 (curve 2). The magnitude of the band gap obtained nanocrystalline cerium dioxide amounted to 3.45 eV (Figure 3). The dependence of the optical density of a solution of methyl violet and ferrous iron in the presence of the thus obtained coated nanocrystalline cerium dioxide from the amount added to the hydrogen peroxide solution is shown in Figure 4 (curve 2). The achievement of the technical result, namely antioxidant activity of the obtained preparation is shown in Figure 5.

Example 3

The method of obtaining implemented similar to that described in Example 1, except that the amount of maltodextrin was 4 g (~0.022 M in terms of glucose).

The particle size of nanocrystalline cerium dioxide was 4 nm, and the hydrodynamic diameter of the particles is covered with a stabilizing shell nanocrystal dioxide Church who I was 7.5 nm (Figure 1). Absorption spectrum obtained Zola in units of molar extinction are shown in figure 2 (curve 3). The magnitude of the band gap obtained nanocrystalline cerium dioxide amounted to 3.55 eV (Figure 3). The dependence of the optical density of a solution of methyl violet and ferrous iron in the presence of the thus obtained coated nanocrystalline cerium dioxide from the amount added to the hydrogen peroxide solution is shown in Figure 4 (curve 3). The achievement of the technical result, namely antioxidant activity of the obtained preparation is shown in Figure 5.

Example 4

The method of obtaining implemented similar to that described in Example 1, except that the amount of maltodextrin was 2.3 g (~0.013 M in terms of glucose).

The particle size of nanocrystalline cerium dioxide was 5 nm, and the hydrodynamic diameter of the particles is covered with a stabilizing shell of nanocrystalline cerium dioxide was 10 nm (Figure 1). Absorption spectrum obtained Zola in units of molar extinction are shown in figure 2 (curve 4). The magnitude of the band gap obtained nanocrystalline cerium dioxide amounted to 3.62 eV (Figure 3). The dependence of the optical density of a solution of methyl violet and ferrous iron in the presence of the thus obtained coated about what accoi nanocrystalline cerium dioxide from the amount added to the hydrogen peroxide solution is shown in Figure 4 (curve 4). The achievement of the technical result, namely antioxidant activity of the obtained preparation is shown in Figure 5.

The obtained particles are covered with a stabilizing shell of nanocrystalline cerium dioxide was analyzed using the methods of dynamic light scattering (in solution) and transmission electron microscopy (Figure 1). It is seen that, depending on the quantity used of the stabilizer, the particle size varies in the range of 2÷5 nm, and their hydrodynamic diameter in the range 6÷10 nm. Dextrose equivalent (DE) and the average degree of polymerization (SP) maltodextrine connected by the relation DE×SP=120, therefore, used maltodextrin with DE-12 consists of amilozid fragments containing 10-12 slices of glucose. Computer simulation of particle size of 2 nm, stable low-molecular oligodactyly connection with aminosol containing an appropriate number of glucose units, shows that depending on the orientation of the molecules of the stabilizer particle size when adsorption is increased by 2÷6 nm (6). The calculation results are in good agreement with the experimental data. Thus, varying amounts of stabilizer, can be targeted to control the size of particles coated with a stabilizing shell of nanocrystalline cerium dioxide.

The size of the particles of cerium dioxide depend on the oxygen stoichiometry and electronic properties of nanocrystalline cerium dioxide [Vchebanov, Aberbach and Averatec, USP 78, (9), 924 (2009)]. Figure 2 shows absorption spectra of the obtained sols (in units of molar extinction). Figure 3 displays the width of the forbidden zone of the obtained particles. Accordingly, oxygen stoichiometry and electronic properties of NDC determine the ability of the particles to participate in redox processes, as well as the ability to inactivate free radicals and protect against oxidative stress.

Antioxidant activity covered with a stabilizing shell of nanocrystalline cerium dioxide by the following studies. Description of the methods and results of the research are listed below.

Inaktivirovanie hydroxyl radical.

Highly reactive short-lived hydroxyl radical (*OH) is one of the most dangerous reactive oxygen species. Hydroxyl radical is easily formed during the Fenton reaction: Fe2++H2O2→Fe3++*OH+OH-.

This radical oxidizes many organic substances, initiates the chain reaction of lipid peroxidation and discolor dyes. For example, the dye methyl violet occurs after the respective reaction:

The speed of bleaching of the dye methyl violet can be registered and education spending in the system hydroxyl radical [Ying Xue, Qingfen Luan, Dan Yang, Xin Yao, Kebin Zhou, J. Phys. Chem. C, 2011, 115 (11), pp. 4433-4438]. Figure 7 shows a set of curves describing the variation of the optical density of the dye solution of methyl violet and salts of Fe2+adding different amounts of hydrogen peroxide.

Materials and methods

Sulphate of iron (II) uranyl, freshly prepared basic solution of 0.2 M hydrogen Peroxide base solution of 0.5 M Methyl violet, basic solution of 50 μm. Powders coated with a stabilizing shell of nanocrystalline cerium dioxide obtained according to the methods described in Examples 1-4.

The execution of the experiment

To 100 ml of a basic dye solution was added 1 ml of a solution of ferric sulfate and record the optical density. Then, under stirring, to this solution was added sequentially a solution of hydrogen peroxide (the total number in the system, 0.1 ml; 0.4 ml; 1.0 ml; 2.0 ml; 3.0 ml etc). After each addition of H2O2recorded the absorption spectrum of a solution.

A portion covered with a stabilizing shell of nanocrystalline cerium dioxide was taken so that regardless of the mass fraction basis of the aqueous substance in the sample, the content of cerium dioxide in it was constant (200 μm). This number was made in 100 ml of basic dye solution and performed the study, as described in the preceding paragraph.

Results and conclusions

The measurement results are shown in Figure 4.

From Figure 4 it is seen that covered with a stabilizing shell nanocrystalline ceria inhibits the decomposition of the dye under the action of hydroxyl radical generated by the Fenton reaction, and depending on the method by which the speed of inaktivirovanie hydroxyl radical sequentially changed. Thus, this invention allows to purposefully get covered with a stabilizing shell nanocrystalline ceria with a given antioxidant properties with respect to the active forms of oxygen.

Protection of mammalian cells from oxidative stress.

Hydrogen peroxide is another common form of active forms of oxygen, including the starting material for the formation of hydroxyl radicals in the biological cell. It is known that hydrogen peroxide causes irreversible cell damage and death by mechanism oxidase.

Materials and methods.

Hydrogen peroxide, 0.1 M solution. Cells mouse fibroblasts (L929). Powders coated with a stabilizing shell nanocryst lifestage dioxide cerium, obtained according to the methods described in Examples 1-4.

The effect on cell viability covered with a stabilizing shell of nanocrystalline cerium dioxide was studied using a reference cell line of mice fibroblasts (L929) from Museum of cell cultures at the Institute of experimental pathology, Oncology and radiobiology. By R.Kavetsky of NAS of Ukraine.

For the formation of a monolayer of cells in the wells of plates (96-well card "Costar, USA) were made in 0.1 ml of a suspension containing 5×105cells/ml, and incubated 24 hours at 37°C in thermostat TC-80 M-2, in an atmosphere with a humidity of 98%, containing 5% CO2. As the environment of growth used a synthetic nutrient medium 199 ("production company", Ukraine), containing 5 to 10% of fetal serum of calves ("Sigma", USA), 25 mm HEPES, 10 mm glutamine, penicillin and streptomycin (100 u/ml each).

Part of a supportive environment consisted of medium 199, 2% fetal serum of calves, 25 mm HEPES, 10 mm glutamine, penicillin and streptomycin (100 u/ml each). For washing the monolayer of cells used medium 199 without serum.

The effect of protection of the studied nuclei coated with a stabilizing shell of nanocrystalline cerium dioxide from oxidative stress was determined from the change in the viability of L929 cells. Each dilution of the analyte tested the Wali three times.

Covered with a stabilizing shell nanocrystalline ceria contributed to formed a monolayer of cells for 24 hours before processing. A portion covered with a stabilizing shell of nanocrystalline cerium dioxide was taken so that regardless of the mass fraction of the main substances in the sample, the content of cerium dioxide in the hole was constant (1 mm).

4 hours after adding hydrogen peroxide in 96-well card with the formed monolayer (final concentration in the well was 0.5 mm) was determined by the number of surviving cells, which was assessed by the intensity of color of the crystal violet cell according to [Medvedev, A.E., Fuchs CENTURIES, Rakhmilevich A.L. A study of the action of immunosuppressive factors from tumor cells on lymphocytes and macrophages in vitro and on the graft-versus-host reaction in mice // Biomed. Sci. - 1990. - V.3, N.1. - P.261-266].

To do this, from the wells was removed the supernatant, and the cells for 15 minutes contributed 0.2% solution of the dye Crystal Violet (Sigma, USA) in 2% ethanol. The dye was removed and the coated monolayer cells were washed with distilled water and dried. The optical density of the stained cells was measured on a spectrophotometer with a vertical beam LabsystemMultiscan (UK) at a wavelength of 540 nm. The percentage of stained cells in the experimental wells was determined by the formula

(Pop-PCP/Pcounter-PCP)*100, where

Pop- performance Opti is eskay density of pilot holes,

Pcounter- the optical density in intact control holes

PCP- the optical density in the wells of the control action of the peroxide.

Statistical processing of data produced in accordance with the recommendations of [Glanz S. biomedical statistics. - M.: "the Practice", 1998. - 459 S., Gubler E.V., Genkin A.A. Application of non-parametric criteria of statistics in biomedical research. - M.: Medicine, 1973. - 142 S.] using complex software patch BIOSTAT 2009 Professional 5.8.1. Numeric data are presented as medians and first and third interquartile interval.

The results and conclusions from the study of antioxidant activity.

The research results are summarized in figure 5. Numbers on the x-axis correspond to the numbers of Examples. The number of living cells without treatment with hydrogen peroxide and nanocrystalline cerium dioxide is taken as 100%. The value "0" corresponds to the control cultures treated only with hydrogen peroxide and a stabilizer (maltodextrin) without the introduction covered with a stabilizing shell of nanocrystalline cerium dioxide. In the latter case there is the absence of any protective actions and death of the cell culture.

From figure it is visible, that is covered with a stabilizing shell nanocrystalline ceria protects cells and mammals from destruction under the action of hydrogen peroxide, moreover, depending on the method by which the effectiveness of protective actions sequentially changed. In some cases (Examples 1 and 2) after processing of nanocrystalline cerium dioxide coated with a stabilizing shell and hydrogen peroxide, the amount of living fibroblasts than untreated culture (values above 100%). That is, there is not only protective, but also stimulating effect of nanocrystalline cerium dioxide obtained according to the proposed method. Thus, this invention allows to purposefully get covered with a stabilizing shell nanocrystalline ceria with specified properties in relation to the protection of biological objects from oxidative stress.

The proposed invention allows to obtain the drug for the protection of biological objects from oxidative stress with high antioxidant activity, with the ability to purposefully vary the size of the particles of cerium dioxide, stable when stored in a dry powdered state, full redispersible resulting product in water or another polar solvent, with the formation of stable non-aggregated Zola with both low and high concentration, and the simplicity of the process execution.

1. The method of obtaining covered stabil shirouma shell nanocrystalline cerium dioxide, characterized by antioxidant activity, which consists in the fact that preparing an aqueous solution of cerium salt and a stabilizer with a molar ratio of cerium and stabilizer 1:1÷4, in terms of the number of monomers in the structure of the stabilizer, wherein the stabilizer is used maltodextrin, to the resulting aqueous solution was dropwise added with stirring an aqueous solution of ammonia and slowly increase the pH of the resulting solution to a value of 7÷8, incubated for 1÷4 h, the obtained colloidal solution of nanoparticles hydroxocobalamine cerium add aqueous ammonia and raise the pH to 11÷12, then incubated for 1÷10 h prior to the formation of a colloidal solution of cerium dioxide, then add an excess of alcohol, selected from a number of: methanol, ethanol, isopropanol, propanol; or ketone selected from the range: dimethylketone, methyl ethyl ketone, diethylketone; and bring to a boil, the precipitate non-aggregated nanoparticles of cerium dioxide coated with a stabilizing shell is separated by decantation or by filtration, washed with 1÷4 times specified alcohol or ketone and dried at a temperature of 50÷80°C until constant weight, and redispersion resulting powder non-aggregated nanoparticles of cerium dioxide coated stabilizing shell in the polar race is varicela leads to the formation of aggregately-resistant Zola.

2. The method according to claim 1, characterized in that salts of cerium use water-soluble cerium salt with a solubility of not less than 6·10-3mol of cerium in 1 l of water.

3. The method according to claim 1, characterized in that the polar solvent used water, dimethylsulfoxide, formamide, ethylene glycol, glycerol.



 

Same patents:

FIELD: electrical engineering.

SUBSTANCE: invention may be used for production of individual crystals of zinc oxide and arrays thereof for application as active elements, material for photocatalytic water treatment, piezoelectric sensors as well as for fundamental physical studies of crystal growth kinetics. Crystals are grown in the air using a continuous action ytterbium fibre laser with yellow metal surface with a layer of multi-walled carbon nanotubes applied thereon treated with such laser radiation with power density equal to approximately 105 W/cm2 during 10 sec. The method enables production of micro- and nanostructured zinc oxide arrays consisting of filamentary crystals, microplates and druses.

EFFECT: invention enables crystals production without special catalysts or crystallisation chambers.

2 dwg

FIELD: chemistry.

SUBSTANCE: fluoride nanoceramic is obtained by thermomechanical treatment of the starting crystalline material made from CaF2-YbF3, at plastic deformation temperature to obtain a workpiece in form of a polycrystalline microstructured substance, which is characterised by crystal grain size of 3-100 mcm and a nanostructure inside the grains, by annealing on air at temperature of not less than 0.5 of the melting point with compaction of the obtained workpiece in a vacuum at pressure of 1-3 tf/cm2 until the end of the deformation process, followed by annealing in an active medium of carbon tetrafluoride at pressure of 800-1200 mmHg. The starting crystalline material used can be a fine powder which has been subjected to heat treatment in carbon tetrafluoride, or a moulded workpiece of crystalline material made from the powder and heat treated in carbon tetrafluoride.

EFFECT: invention enables to obtain a fluoride nanoceramic with high degree of purity and high uniformity of the structure of said optical material.

4 cl, 3 ex

FIELD: chemistry.

SUBSTANCE: fluoride nanoceramic is obtained by thermomechanical treatment of the starting crystalline material made from CaF2-YbF3, at plastic deformation temperature to obtain a workpiece in form of a polycrystalline microstructured substance, which is characterised by crystal grain size of 3-100 mcm and a nanostructure inside the grains, by annealing on air at temperature of not less than 0.5 of the melting point with compaction of the obtained workpiece in a vacuum at pressure of 1-3 tf/cm2 until the end of the deformation process, followed by annealing in an active medium of carbon tetrafluoride at pressure of 800-1200 mmHg. The starting crystalline material used can be a fine powder which has been subjected to heat treatment in carbon tetrafluoride, or a moulded workpiece of crystalline material made from the powder and heat treated in carbon tetrafluoride.

EFFECT: invention enables to obtain a fluoride nanoceramic with high degree of purity and high uniformity of the structure of said optical material.

4 cl, 3 ex

FIELD: chemistry.

SUBSTANCE: aqueous solution of quantum dots based on cadmium selenide coated with mercapto acids is stabilised by adding sodium sulphite until achieving its concentration of 0.02-0.2 mol/l in the solution.

EFFECT: high stability of aqueous solution of quantum dots while preserving luminescence brightness, hydrodynamic diameter and active groups of the quantum dots.

2 dwg

FIELD: chemistry.

SUBSTANCE: aqueous solution of quantum dots based on cadmium selenide coated with mercapto acids is stabilised by adding sodium sulphite until achieving its concentration of 0.02-0.2 mol/l in the solution.

EFFECT: high stability of aqueous solution of quantum dots while preserving luminescence brightness, hydrodynamic diameter and active groups of the quantum dots.

2 dwg

FIELD: chemistry.

SUBSTANCE: invention relates to inorganic chemistry and can be used in making catalyst supports, filters and electronic materials. The starting materials used are powdered silicon and detonation synthesis diamonds, which are mixed in weight ratio silicon:diamond from 2:1 to 2.4:1. The powdered mixture is then moulded into a workpiece of the required article and then heat treated at temperature of 1200-1500°C in a vacuum or inert medium.

EFFECT: invention enables to obtain nanostructured silicon carbide in form of articles having high porosity - 55-65 vol. % and simultaneously sufficient strength, wherein the size of crystalline regions is in the range of 10-50 nm.

FIELD: nanotechnologies.

SUBSTANCE: invention relates to technology of nanoparticles production. The method is proposed to produce hollow nanoparticles, which consists in application of a silicon carbide coating onto a surface of carbon nanotubes by pyrolysis of methyl silane, afterwards carbon nanotubes are removed by burning.

EFFECT: increased heat resistance of hollow nanoparticles and reduced duration of a production cycle in process of their production.

1 cl, 3 dwg

FIELD: chemistry.

SUBSTANCE: invention can be used in plasma chemistry and pharmacology. A vapour-liquid plasmatron is used to form a plasma jet from alcohol or aqueous solution thereof. The plasma jet is fed into a volume of water by dipping the nozzle of the plasmatron 10-20 mm into the water perpendicular to the water surface.

EFFECT: obtaining nanodiamonds with size of 20-40 nm in a continuous process.

3 dwg

FIELD: chemistry.

SUBSTANCE: invention relates to chemical industry and medicine and can be used in producing concrete plasticisers, microbiocides with anti-HIV properties which are not cytotoxic and modifiers of epoxy composites. Fullerene-polysulphonic acid is a water- and alcohol-soluble product of reaction of coal tar and coal tar pitch with sulphuric acid with concentration of at least 80%. Unreacted sulphuric acid formed when treating sulphonic acid and unreacted aromatic hydrocarbons are successively washed off with toluene, carbon tetrachloride and acetone to pH 6.5-7.0. Fullerene-polysulphonic acid is then extracted with ethyl alcohol in Soxhlet apparatus. Ethyl alcohol is distilled from the alcohol solution of fullerene-polysulphonic acid to dry residue.

EFFECT: invention enables to obtain fullerene-polysulphonic acid using a simple method using cheap and readily available reactants.

7 cl, 2 dwg, 2 ex

FIELD: chemistry.

SUBSTANCE: invention relates to chemistry and nanotechnology. The method of producing fullerene C84 involves treating coal tar or coal tar pitch with sulphuric acid, washing off unreacted sulphuric acid, the formed sulphonic acids and hydroxyl compounds with water, and the unreacted aromatic hydrocarbons successively with carbon tetrachloride and acetone. Fullerene is then extracted with toluene or ortho-xylene and the toluene or ortho-xylene are distilled from the obtained extract until achieving a moist state of fullerene residue which is dried at temperature of 60-70°C. The obtained fullerene C84 is soluble in benzene, toluene and ortho-xylene.

EFFECT: invention enables to obtain fullerene C84 using a cheap and low power consumption method.

11 cl, 3 ex

FIELD: medicine.

SUBSTANCE: invention relates to method of obtaining beta-lactam antibiotics in for of polymer complex gels with silver ions, which includes preparation of solutions of sodium salts of antibiotics of 0.1-1.0 mol/l concentration, solution of silver nitrate of 0.1-1.0 mol/l concentration, mixing observing ratio antibiotic:silver not less than 1 and not higher than 9, obtained solutions at temperature 10÷40°C with formation of dispersion of white colour, keeping dispersion without mixing until its decolouration with formation of colourless transparent gels, characterised by the fact that as antibiotic used are oxacillin derivatives, solutions of sodium salts of antibiotic contain 50-90 volume percent of water and 10-50 volume percent of organic solvent.

EFFECT: gel is thermally stable, ensures higher bactericidal and antibacterial activity and is prepared without additional polymer thickening agents.

11 dwg, 4 tbl, 3 ex

FIELD: measurement equipment.

SUBSTANCE: first, the appropriate planar integral circuit is created; for that purpose, contact platforms are formed on reverse side of the base; then, the first conducting layer is created; after that, circuit of a sensitive element of the converter is formed and a sacrificial layer is removed, and the method is completed with formation of the converter housing and formation of whiskers. Besides, at formation of the converter housing in a closed volume of the housing, gas medium with specified parameters is created, and formation of a whisker is performed due to supply of pulses of the specified shape to electrodes. As specified parameters of the medium, for example, pressure and/or dew point are used. Pulses in the form of half-period of sine wave of the specified amplitude and frequency are supplied to electrodes.

EFFECT: unification of different types of sensors with autoelectronic emission and reduction of standard sizes of different sensors.

4 cl, 12 dwg

FIELD: measurement equipment.

SUBSTANCE: in pressure measurement method using a pressure strain gauge based on nano- and microelectromechanical system (NaMEMS), in the measurement mode, value of measured pressure Pi is calculated by means of biharmonic spline interpolation at check points based on column-vector W(P0, Uiz, Upl) saved at calibration stage by the following formula: Pi=GT×W, where GT - transposed column-vector G; "x" means matrix product. Calibration for pressure measurement is performed as per the method consisting in recording of stresses of measuring Uiz and feeding Upl diagonals of bridge measuring circuit and recording to a read-only memory of sensor column-vector W, which is calculated by the following formula: W=g-1×P, where P - column-vector of reference pressure values at check points; g - matrix, the diagonal elements of which are equal to zero, and the rest components are calculated in a certain way. Pressure sensor based on NaMEMS, implementing the proposed measurement and calibration methods, involves a calculation device that includes a conversion unit of ADC code to numerical value of stress, a calculation unit of numerical pressure value; at that, in the calculation unit of numerical value of pressure, the measured pressure Pi is determined by the formula Pi=GT×W.

EFFECT: improving measurement accuracy of pressure and processibility.

3 cl, 3 dwg

FIELD: electrical engineering.

SUBSTANCE: invention may be used for production of individual crystals of zinc oxide and arrays thereof for application as active elements, material for photocatalytic water treatment, piezoelectric sensors as well as for fundamental physical studies of crystal growth kinetics. Crystals are grown in the air using a continuous action ytterbium fibre laser with yellow metal surface with a layer of multi-walled carbon nanotubes applied thereon treated with such laser radiation with power density equal to approximately 105 W/cm2 during 10 sec. The method enables production of micro- and nanostructured zinc oxide arrays consisting of filamentary crystals, microplates and druses.

EFFECT: invention enables crystals production without special catalysts or crystallisation chambers.

2 dwg

FIELD: chemistry.

SUBSTANCE: fluoride nanoceramic is obtained by thermomechanical treatment of the starting crystalline material made from CaF2-YbF3, at plastic deformation temperature to obtain a workpiece in form of a polycrystalline microstructured substance, which is characterised by crystal grain size of 3-100 mcm and a nanostructure inside the grains, by annealing on air at temperature of not less than 0.5 of the melting point with compaction of the obtained workpiece in a vacuum at pressure of 1-3 tf/cm2 until the end of the deformation process, followed by annealing in an active medium of carbon tetrafluoride at pressure of 800-1200 mmHg. The starting crystalline material used can be a fine powder which has been subjected to heat treatment in carbon tetrafluoride, or a moulded workpiece of crystalline material made from the powder and heat treated in carbon tetrafluoride.

EFFECT: invention enables to obtain a fluoride nanoceramic with high degree of purity and high uniformity of the structure of said optical material.

4 cl, 3 ex

FIELD: biotechnologies.

SUBSTANCE: invention is designed for extraction and treatment of DNA and RNA from biological samples - samples with purity suitable for their subsequent analysis by the method of polymerase chain reaction in real time (Real Time PCR). The device comprises a body, test tubes for placement of nucleic acids adsorbed on magnetic particles in each of them and a pestle magnetised perpendicularly. The body is made of non-magnetic material. In the body there are the following components installed as capable to generate rotary non-uniform magnetic field: a horizontally arranged shaft and magnetic cylinders. The shaft comprises blocks of four crossed permanent magnets that repeat along the longitudinal axis of the shaft. The position of each magnet is determined by the angle of rotation of identical poles relative to the first magnet, the second magnet is turned by 90 degrees, the third one - by 270 degrees, the fourth magnet - by 180 degrees. Magnetic cylinders are arranged at both sides of the horizontal shaft. Each of the magnetic cylinders is magnetised perpendicularly as capable of rotation only around this axis, and is arranged in parallel to the test tube wall. Each pair of magnetic cylinders at different sides of the longitudinal axis of the horizontal shaft is arranged as capable of interaction with two crossed permanent magnets of the horizontal shaft, besides, the horizontal shaft is made and placed in the body so that its axis is perpendicular to the axes of magnetic cylinders. The number of pairs of crossed permanent magnets on the shaft is equal to the number of magnetic cylinders arranged at one side from the longitudinal axis of the shaft, at the same time test tubes are arranged in the body in magnetic non-uniform rotary field.

EFFECT: increased efficiency of wear of a coil of magnetic particles entangled with nucleic acids and production of samples with purity suitable for their subsequent analysis by the method of polymerase reaction.

5 dwg

FIELD: chemistry.

SUBSTANCE: aqueous solution of quantum dots based on cadmium selenide coated with mercapto acids is stabilised by adding sodium sulphite until achieving its concentration of 0.02-0.2 mol/l in the solution.

EFFECT: high stability of aqueous solution of quantum dots while preserving luminescence brightness, hydrodynamic diameter and active groups of the quantum dots.

2 dwg

FIELD: chemistry.

SUBSTANCE: invention relates to inorganic chemistry and can be used in making catalyst supports, filters and electronic materials. The starting materials used are powdered silicon and detonation synthesis diamonds, which are mixed in weight ratio silicon:diamond from 2:1 to 2.4:1. The powdered mixture is then moulded into a workpiece of the required article and then heat treated at temperature of 1200-1500°C in a vacuum or inert medium.

EFFECT: invention enables to obtain nanostructured silicon carbide in form of articles having high porosity - 55-65 vol. % and simultaneously sufficient strength, wherein the size of crystalline regions is in the range of 10-50 nm.

FIELD: nanotechnologies.

SUBSTANCE: invention relates to technology of nanoparticles production. The method is proposed to produce hollow nanoparticles, which consists in application of a silicon carbide coating onto a surface of carbon nanotubes by pyrolysis of methyl silane, afterwards carbon nanotubes are removed by burning.

EFFECT: increased heat resistance of hollow nanoparticles and reduced duration of a production cycle in process of their production.

1 cl, 3 dwg

FIELD: chemistry.

SUBSTANCE: invention can be used in plasma chemistry and pharmacology. A vapour-liquid plasmatron is used to form a plasma jet from alcohol or aqueous solution thereof. The plasma jet is fed into a volume of water by dipping the nozzle of the plasmatron 10-20 mm into the water perpendicular to the water surface.

EFFECT: obtaining nanodiamonds with size of 20-40 nm in a continuous process.

3 dwg

FIELD: medicine.

SUBSTANCE: group of inventions refers to medicine, namely to radiation biology, and may be used to treat a cadmium radiation injury and to produce a preparation for the cadmium radiation injury. Treating the cadmium radiation injury is ensured by the single subcutaneous administration of a composition consisting of anti-radiation therapeutic globulin and a suspension-forming fraction of bentonite, in a dose of 20-25 mg/kg of body weight that is followed by three more administrations 24, 48, 96 hours later if observing chronic heavy metal administration. The preparation is produced by dissolving the suspension-forming fraction of bentonite in anti-radiation therapeutic globulin in ratio 0.3:99.7 respectively while constantly stirred, and a solid concentration is reduced to 10%; the preparation is sterilised by filtration and then bottled 250-300 cm3 each to be closed tightly and kept at temperature 4-6°C.

EFFECT: method enables a consistent process of the preparation with enhanced therapeutic and decorporating effect ensured by the targeted delivery as a single agent with simplifying the use conditions.

3 cl, 2 tbl

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