RU2659949C1 - Method for preparing a preparation based on magnetic nanoparticles (mnch) of iron oxide for mrt-diagnosis of neoplasms - Google Patents

Abstract

FIELD: pharmaceuticals.

SUBSTANCE: invention relates to chemical and pharmaceutical industry and is a method for preparing a preparation for MRI diagnosis of tumor diseases, comprising preparation of a solution of iron (III) acetylacetonate in benzyl alcohol with a concentration of 75–200 g/l, followed by heating in an inert gas stream to boiling point of benzyl alcohol for 4–8 hours and boiling the solution for 30 minutes to 4 hours to obtain a suspension, after which the suspension is cooled, washed with a polar organic solvent to obtain iron oxide nanoparticles Fe₃O₄, which are then coated with human serum albumin and/or bovine serum albumin, and the obtained coating is stabilized by intermolecular cross-linking with glutaraldehyde.

EFFECT: invention provides magnetic nanoparticles with an initial hydrodynamic size (up to stabilization of HSA/BSA) to 20 nm, and nanoparticles stabilized by HSA/BSA up to 50 nm in size but due to change in the boiling point of the iron (III) acetylacetonate solution in benzyl alcohol in preparation of the original magnetic nanoparticles.

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Description

Technical field

The invention relates to chemistry, in particular to a method for producing magnetic nanoparticles of iron oxide, which can be used in medicine as a contrast agent for magnetic resonance imaging (MRI) studies, including for neoplasms of various nature

State of the art

For more accurate diagnosis of various abnormalities using MRI, various contrast agents are widely used, which can be divided into two main classes, the first of which includes T1 contrast agents, which are paramagnetic metal ions containing a large number of unpaired electrons (Gd3 +, Eu3 +, Cr3 + , Mn2 +, Fe3 +), and the second - T2-contrast agents, represented by magnetic nanoparticles of iron oxide, stabilized by various biocompatible coatings.

The claimed invention is directed to the production of magnetic nanoparticles (MNPs) of iron oxide and a T2-contrast drug based on them, stabilized with human serum albumin (BSA) and / or bovine serum albumin (BSA) with a nanoparticle (or globule) size of up to 50 nm. T2-contrast agents are negative contrasts - their accumulation lowers the signal intensity in the modes, weighted by the uniformity of the magnetic field in an MRI study. It is significant that T2-contrast agents, having orders of magnitude higher magnetic susceptibility due to the iron oxide nanocrystals included in them, have T2-relaxivity 50-100 times higher than T1-contrast agents, which reduces the necessary dose for effective contrasting.

Today, the main methods for producing magnetic nanoparticles are the thermal decomposition of organometallic precursors in high-boiling organic solvents in the presence of surface active substances (surfactants) (method 1 - A.N. Lu, EL Salabas, F. Schuth, Magnetic nanoparticles: Synthesis, protection, functionalization , and application, Angew. Chemie - Int. Ed. 46 (2007) 1222-1244. doi: 10.1002 / anie.200602866.) and coprecipitation of metal salts in stoichiometric ratios (method 2 - A.K. Gupta, M. Gupta, Synthesis and surface engineering of iron oxide nanoparticles for biomedical applications, Biomaterials. 26 (2005) 3995-4021. Doi: 10.1016 / j.biomaterials. 2004.10.012). Method 1 allows one to obtain monodisperse nanoparticles, however, the introduction of surfactants makes the obtained nanoparticles hydrophobic, which prevents their dissolution in an aqueous medium and does not allow for use in the body. Method 2 allows you to get hydrophilic nanoparticles, however, a feature of the method is to obtain a mixture of heterogeneous in size nanoparticles, which does not allow their use in the future because of the inconstancy of the composition and size distribution.

There are also other methods, such as pyrolysis, microwave synthesis, sonochemical synthesis, however, they do not allow to obtain iron oxide nanoparticles with the required properties suitable for use in MRI diagnostics.

When using magnetic nanoparticles in clinical practice, biocompatible, non-toxic, biodegradable components are introduced into the composition of preparations based on them to give the drug stability and reduce toxicity.

Resovist®, manufactured by Bayer Schering Pharma AG, which is a magnetic nanoparticle coated with dextran larger than 100 nm, is known in the art.

However, this drug is characterized by very short circulation times in the bloodstream (2-5 min), associated with a nanoparticle size of more than 100 nm, as well as accumulation only in the liver, which significantly limits its use in the diagnosis of tumor diseases of other organs.

The prior art also discloses a magnetic nanocomposite for a contrast agent, a contrast agent and a drug delivery agent for diagnosis and treatment (patent application US 20130045160 A1 of 04.17.2013). This source describes the coating of nanoparticles with amphiphilic compounds, including albumin, to obtain a magnetic nanocomposite consisting of one or more magnetic nanoparticles distributed in the hydrophobic domain of the amphiphilic compound, and a shell containing the hydrophilic domain of the amphiphilic compound, while the hydrophobic domain is associated with the surface of magnetic nanoparticles through physical bonding, rather than chemical.

The disadvantage is the fact that physical bonds are orders of magnitude weaker than chemical bonds, which leads to instability of these coatings. In addition, if it enters the bloodstream, in case of physical immobilization due to hydrophobic interactions, the shell of the amphiphilic compound will be replaced by plasma proteins, and the amphiphilic polymers themselves can also be sorbed on the surface of blood cells, which will lead to a change in the pharmacokinetics of the drug and impairment of its delivery to the tumor hearth.

The prior art also knows the solution presented in the application for the invention US 20080206146 A1 dated 09.11.2007, describing magnetic nanoparticles, a method for their preparation and use. The document discloses magnetic nanoparticles containing a functional group that provides differential binding to brain tissue, blood vessels and bone tissue, intended for use as a diagnostic tool in MRI studies and as a means of drug delivery. The functionalized magnetic nanoparticle is encapsulated in an albumin matrix. Encapsulation in albumin involves the following steps: 200 mg of human serum albumin (HSA) is dissolved in 2.0 ml of water containing magnetic nanoparticles (MNP, for example, magnetite particles). The Ph of the solution is increased to 8.4 with constant stirring by adding dropwise a 0.01 M and 0.1 M NaOH solution. Then, with constant stirring, 8.0 ml of ethanol was added dropwise, thereby obtaining a 10% HSA solution. After adding ethanol, 235 μl of an 8% glutaraldehyde solution is added. After 24 hours, the resulting nanoparticles are purified by three-fold centrifugation (16,100 g, 8 min) and repeated resuspension in water in an ultrasonic bath. Functionalized magnetic nanoparticles coated with HSA have an average diameter of 60 nm to 990 nm, depending on the pH of the preparation and the particles used.

However, the preparation of these nanoparticles is characterized by significant polydispersity of the nanoparticles (size dispersion from 60 to 990 nm), which limits its use in practice, since large nanoparticles have a significantly shorter half-life, which does not allow them to be effectively delivered to the tumor site.

The difference between the proposed solution and the known one consists in using thermal decomposition of iron acetylacetonate to produce monodispersed magnetic nanoparticles with an initial hydrodynamic size of up to 20 nm, mainly up to 12 nm, capable of dissolving in water with subsequent immobilization of BSA / BSA, stabilization with glutaraldehyde and the preparation of preparations containing MNP with a hydrodynamic particle size of less than 50 nm, which eliminates the disadvantages characteristic of these analogues.

The prior art method for producing magnetic nanoparticles (Hilda TR Wiogo, May Lim, Volga Bulmus, Jimmy Yun, and Rose Amal Stabilization of Magnetic Iron Oxide Nanoparticles in Biological Media by Fetal Bovine Serum (FBS) // Langmuir 2011, Vol. 27, No. 2, P. 843-850.) Having carboxyl groups on the surface by mixing with fetal calf serum. It is indicated that such particles have an average hydrodynamic diameter of 180 nm and retain it for 16 hours of incubation in RPMI nutrient medium. It has also been shown that protein immobilization occurs non-covalently. However, the large diameter of the nanoparticles (180 nm) can impede their diffusion through the blood vessels to the tumor and thereby reduce their effectiveness as a contrast agent. Non-covalent surface modification can lead to desorption of the coating when introduced into the bloodstream and, accordingly, to the loss of all components attached to the coating. In addition, in this article there is no data on the value of T2-relaxivity necessary to assess the effectiveness of the obtained contrast agent for MRI diagnostics.

The closest analogue of the proposed invention is a contrast agent for conducting an MRI study, a method for its production and a method for MRI diagnostics of glioblastoma multiforme (patent RU 2530762 of 12/14/2012; Abakumov MA, et al. Nanomedicine. May 2015, Volume 11, Issue 4, pp. 825-833). According to these publications, thermal decomposition of iron (III) acetylacetonate in benzyl alcohol was used to obtain magnetic nanoparticles. The synthesis of MNPs was carried out in an inert gas stream with constant stirring of the reaction mixture and slow heating to a temperature of 383 K, after which it was held for 1 h at this temperature to evaporate water from the solution, then the temperature of the reaction mixture was raised to a speed of 25 deg / h to 473 K, which a total of 9 hours, and kept at this temperature for 40 hours. After 40 hours, the reaction mixture was slowly cooled to room temperature, 90 ml of anhydrous acetone was added and MNP was separated by centrifugation at 2000 g for 10 min. This precipitate was washed twice with excess acetone and then dried on a rotary evaporator until acetone was completely removed. The diameter of the obtained nanoparticles was, according to transmission electron microscopy, (14 ± 4) nm. At the next stage of the technological process, aqueous colloidal solutions of iron oxide nanoparticles were stabilized, for which bovine serum albumin (BSA) was used. For this, 5 ml of distilled water was added to 10 mg of particles and the pH was adjusted to 11 with 1 M NaOH solution. Then, the resulting dispersion was sonicated for 10 min, and 40 mg of polymer dissolved in 5 ml of water was added thereto. The resulting mixture was incubated for 4 hours at room temperature with constant stirring, then dialyzed against distilled water and 500 μl of 1M NaOH was added, and then 2.3 ml of a 25% aqueous solution of glutaraldehyde was added dropwise with stirring. The resulting mixture was incubated with stirring for 15 minutes, and then 500 μl of 3 M glycine with a pH of 9.2 was added to bind unreacted aldehyde groups. To the resulting solution was added 1 ml of a solution of sodium borohydride in PBS at a concentration of 2 mg / ml. Then incubated for 60 minutes. To separate the BSA coated MNPs (MNP-BSA) from excess protein, the NP solution was passed through cellulose centrifuge filters with a pore diameter of 100 kDa. Then, the resulting precipitate was resuspended in water and again subjected to filtration. The procedure was repeated until the protein completely disappeared in the washing liquid. The protein was purified from molecular crosslinking products by gel filtration on a Sepharose CL-6B support (column heights and diameters of 50 and 2.5 cm, respectively, flow rate 0.7 ml / min). At the next stage, conjugates of magnetic nanoparticles with monoclonal antibodies to vascular endothelial growth factor antibodies having a hydrodynamic diameter of less than 150 nm were obtained, which were further used as a contrast agent for MRI diagnostics of glioblastoma multiforme.

According to the method according to the patent RU 2530762, an MRI study was carried out before and after intravenous administration of the obtained contrast agent, while an MRI study was carried out in a mode providing obtaining a magnetic susceptibility-weighted image of the test site, and the conclusion about the presence of glioblastoma multiforme is made according to the results of MP- comparison images before and after the introduction of a contrast medium according to areas of diminishing image brightness on MR images.

The known method improves the reliability and informativeness of diagnosis by increasing the contrast of the areas corresponding to the tissues of multiform glioblastoma, its vessels and foci of neoangiogenesis in MP images. However, the obtained nanoparticles had a hydrodynamic diameter of 92 nm. Thus, the disadvantage of the described method is the impossibility of obtaining nanoparticles with an HSA coating of size less than 50 nm due to the use of long boiling in the technological process (for 40 hours at a temperature of 200 ° C). During boiling, the processes of chemical aging of the surface of nanoparticles occur (the conversion of OH groups to oxo-bridges), which leads to a decrease in the number of charged groups on the surface of nanoparticles, which leads to a weakening of the stabilization of electrostatic repulsion and a decrease in the charge density necessary for protein sorption. In addition, the processes of chemical aging lead to an increase in the initial size of the nanoparticles, which negatively affects their stability during resuspension in an aqueous medium. In general, both of these processes lead to the fact that nanoparticles obtained after prolonged heating are substantially less stable in the aquatic environment and tend to aggregate, while the aggregation of the initial magnetic nuclei also leads to an increase in the size of the final nanoparticles (globules) already coated with HSA.

Disclosure of invention

The problem to be solved in the proposed invention is the development of a method for producing magnetic nanoparticles stabilized with serum albumin, suitable for use as MRI contrast agents for imaging tumors.

The technical result of the invention is the ability to obtain magnetic nanoparticles with an initial hydrodynamic size (before stabilization of BSA / BSA) up to 20 nm, and nanoparticles (globules) stabilized by HSA / BSA, up to 50 nm in size, by changing the boiling mode of the solution of iron (III) acetylacetonate in benzyl alcohol, upon receipt of the initial MNPs, no more than 4 hours. When boiled for more than 4 hours, irreversible aging processes occur on the surface of the MNPs, preventing them from dissolving in water and sorption of HSA onto their surface.

The drug obtained by the claimed method provides an increase in the efficiency of MRI studies of neoplasms by improving the penetration of the drug into the tumor site when it is introduced in the MRI process. In this case, it becomes possible to effectively penetrate through the pores of the defective vessels of the tumor tissue.

The technical result of the invention is achieved by the fact that the method of obtaining a preparation for MRI diagnosis of tumor diseases involves the preparation of a solution of iron (III) acetylacetonate in benzyl alcohol with a concentration of 75-200 g / l, followed by heating in an inert gas stream to the boiling point of benzyl alcohol for 4-8 hours and boiling the solution from 30 min to 4 hours to obtain a suspension, after which the suspension is cooled, washed with a polar organic solvent, to obtain iron oxide nanoparticles Fe ₃ O ₄ , which are then covered with human serum albumin and / or bovine serum albumin, and the resulting coating is stabilized by intermolecular crosslinking with glutaraldehyde.

To coat nanoparticles with human serum albumin and / or bovine serum albumin, nanoparticles are dissolved in water with a pH of 10-11 to obtain a concentration of 2-8 mg / ml, BSA and / or HSA are added to the resulting solution in the form of an aqueous solution with a concentration of the mentioned BSA and / or HSA 4-16 mg / ml in a ratio of 1: 1 by volume, followed by dialysis of the resulting solution.

After stabilization of the coating by intermolecular crosslinking, an additional purification of reaction by-products by ultrafiltration and sterilization by filtration is carried out. In this case, ultrafiltration on filters with a pore diameter of 100-300 kDa is possible.

In a preferred embodiment of the invention, heating is carried out at a constant rate.

As a polar organic solvent, water-soluble aliphatic monohydric alcohols, acetones, ketones, nitriles can be used. In this case, ethanol, methyl alcohol propyl alcohol can be used as water-soluble aliphatic monohydric alcohol, acetone, butanone-2 as ketones, acetonitrile as nitriles.

Washing is carried out until the disappearance of traces of benzyl alcohol. In a particular embodiment of the invention, the washing is carried out in volumes equal to at least the volume of the suspension, at least three times. Washing can also be carried out by centrifugation.

Obtained by the claimed method, the drug can be presented in the form of a solution or lyophilisate.

For an MRI diagnosis of tumor diseases, an MRI scan of the object (human or animal) is carried out prior to administration of the drug in modes having T2 and T2 * weight, then intravenous administration of the drug in an amount of 1-10 mg / kg of the body weight of the object by iron content and MRI studies of the object in the modes with T2 and T2 * weightedness within a period not later than 2 hours after drug administration followed by a comparison of changes in the intensities of the studied area on MRI images obtained before and after conducting prep arata. When hypointensive areas are identified, a conclusion is made about the presence of a tumor disease with the definition of their boundaries.

The inventive method allows to reduce the time to obtain both directly MNPs and a contrast preparation - MNPs with albumin, characterized by the presence of aggregates (NPs) of less than 50 nm in it. Thus, a simplification of the technology for producing a contrast preparation is provided, including and due to the exclusion of the gel filtration stage (when compared with prototype technologies). The inventive method for producing a contrast preparation takes from 6, 5 to 10 hours, known methods - 46 hours. In addition, when compared with technologies of other authors, in their case, the formation of a stable mixture of albumin with MNP does not occur due to chemical bonds. The advantage is that the claimed drug is excreted through the kidneys within a few hours after administration, while other drugs accumulate in the liver (Majumdar S, Zoghbi SS, Gore JC. Pharmacokinetics of superparamagnetic iron-oxide MR contrast agents in the rat. Invest Radiol . 1990; 25: 771-777), providing a reduction in side effects from the administration of the drug. In addition, the claimed drug is stably stored throughout the year as a lyophilisate and resuspended while maintaining parameters.

Brief Description of the Drawings

The invention is illustrated by drawings, where in FIG. 1 shows a micrograph of nanoparticles of iron oxide obtained by the claimed method, while the micrograph is obtained using a transmission electron microscope; in FIG. 2 presents the results of measuring the hydrodynamic dimensions of the MNP obtained after 30 min of boiling in benzyl alcohol; in FIG. 3 - the results of measuring the hydrodynamic dimensions of the MNP obtained after 1 h of boiling in benzyl alcohol; in FIG. 4 - measurement results of the hydrodynamic dimensions of the MNP obtained after 4 hours of boiling in benzyl alcohol; in FIG. 5 - results of measuring the hydrodynamic dimensions of the MNP obtained after 20 hours of boiling in benzyl alcohol; in FIG. 6 - MRI images of a brain tumor of rat C6 before (A) and 5 minutes after (B) administration of a preparation based on MNP of iron oxide obtained by the claimed method; in FIG. 7 - MRI images of the mucous membrane of liver cancer of the RS-1 before (A) and 5 minutes after (B) the introduction of the drug based on MNP of iron oxide obtained by the claimed method; in FIG. 8 - MRI image of adenocarcinoma of the mammary gland of the mouse before (A) and 5 minutes after (B) the introduction of the drug based on MNP of iron oxide obtained by the claimed method.

The implementation of the invention

The method of producing iron oxide nanoparticles of Fe ₃ O _{4 is} preferably used as a contrast medium in MRI diagnosis of tumor diseases, as follows.

Below is a detailed description of the invention with specific parameters of the quantitative content of the components and parameters of the process of obtaining the MNP and preparation for MRI diagnostics, however, this presentation does not limit the claimed invention with the indicated values / parameters, but only demonstrates the possibility of its implementation to achieve the stated result.

For synthesis, a solution of iron (III) acetylacetonate in benzyl alcohol with a concentration of 75-200 g / l is prepared. Then a solution of iron (III) acetylacetonate in benzyl alcohol with a concentration of 75-200 g / l in an inert gas stream is heated to the boiling point of the solvent for 4-8 hours, preferably 6 hours and further boiling for 30 minutes - 4 hours. Reaction mixture cooled to room temperature, centrifuged to separate the nanoparticle precipitate from the solution, washed with acetone / ethanol, or methyl alcohol, or propyl alcohol / acetonitrile, and after evaporation of the solvent, magnetic iron oxide nanoparticles are obtained.

Magnetic nanoparticles (80 ± 20 mg) are placed in 20 ml of distilled water, 500 μl of 1 M NaOH are added to create an alkaline medium, and the solution is stirred until the precipitate disappears. After this, a solution of human serum albumin / bovine serum albumin (160 ± 50 mg in 20 ml of water) is added. The resulting mixture was stirred for 15 minutes, after which large aggregates of particles were removed by filtration on syringe filters with a pore diameter of 0.22-0.45 μm. Then, 500-1500 μl of a 25% aqueous solution of glutaraldehyde are added to the mixture. The reaction mixture is incubated for 10-60 minutes with constant stirring, after which 0.5-2 ml of a 3 M solution of glycine is added to it and stirred for another 30 minutes for 2 hours. After this, 0.5-4 ml of NaBH4 solution (10 mg / ml), and incubated for 30 min - 4 hours

Excess low molecular weight substances and free HSA molecules are removed by washing with phosphate-saline buffer on centrifuge filters with a transmission edge of 100-1000 kDa for globular proteins.

Thus obtained nanoparticles are able to dissolve in distilled water at pH 10-11, and when coated with human serum albumin, they form nanoparticles consisting of crystalline magnetic cores of iron oxide and a shell of human serum albumin with a size of not more than 40 nm.

The experiments are performed on animals with an experimental model of glioma or another tumor. To visualize gliomas, animals are placed in an MP scanner and an MP scan is performed in T2 or T2 * weighted mode. Then the animal is injected intravenously with a solution of MNS stabilized by HSA. The entered dose of MNP in terms of iron concentration is 12010 mg / kg. MRI scans were performed on a Bruker Clinscan 7T scanner. Then, an MRI scan is performed in T2 or T2 * weighted mode for 0-24 hours after intravenous administration.

The advantage of using nanoparticles with a diameter of less than 50 nm in MRI diagnostics is due to the fact that with a decrease in the size of the nanoparticles (globules), their circulation time in the blood increases significantly, which allows to increase the imaging efficiency without resorting to increasing the dose. In addition, tumor vessels have the effect of increased permeability and retention due to overexpression of pro-angiogenic factors leading to accelerated growth of the vascular network. In this case, the pores arising in the vessels have a diameter of 50-200 nm, and only nanoparticles with a diameter of less than 50 nm are able to effectively penetrate through such pores into the tumor tissue.

Example 1. Synthesis of magnetic nanoparticles of Fe ₃ O ₄

10.5 g of iron (III) acetylacetonate and 220 ml of benzyl alcohol are heated with stirring in a glass flask to a temperature of 50-70 ° C for 1 hour. After that, the heating rate is 25 ° C / h. 30 minutes after the temperature of the reaction mixture reaches the boiling point, heating is stopped. 90 ml of acetone was added to the reaction mixture cooled to room temperature, and nanoparticles were precipitated by centrifugation at 900 g for 10 min.

Example 2-5. Synthesis of magnetic nanoparticles of Fe ₃ O ₄

The synthesis of MNPs is carried out according to the method described in example 1, except that instead of acetone, methyl alcohol, ethyl alcohol, propyl alcohol, butanone-2, acetonitrile are added.

Example 6. Synthesis of magnetic nanoparticles of Fe ₃ O ₄ coated with BSA / BSA

To 20 mg of magnetic nanoparticles of iron oxide add 5 ml of distilled water, adjust the pH to 11 and mix until complete dissolution. Then add 5 ml of a solution of BSA / HSA in water with a concentration of 8 mg / ml at the same pH value. The resulting mixture was incubated for 15 minutes at room temperature with constant stirring, then filtered (filter pore diameter 0.2-0.5 μm). To 20 ml of the resulting solution with a protein and Fe concentration of 4 mg / ml and 2 mg / ml, respectively, 250 μl of 1M alkali are added, and then 230 μl of a 25% aqueous solution of glutaraldehyde are added dropwise with stirring. The resulting mixture was incubated with stirring for 15 min, and then 250 μl of a 3M aqueous solution of glycine with a pH of 9.2 were added and incubated for 1 h. Then 332 μl of a solution of sodium borohydride in 10 mg / ml phosphate-buffered saline was added to the solution and incubated 2 h, after which the reaction mixture was washed with buffer and the concentration of iron was measured.

Example 7. MRI imaging of an experimental rat brain tumor C6

The experiments are performed on animals with an experimental model of C6 glioma of another tumor. To visualize gliomas, animals are placed in an MP scanner and an MP scan is performed in T2 * weighted mode with the following parameters: SWI mode: TE / TR = 19/50 ms, cut thickness 0.5 mm, FOV = 30 mm, resolution 256/176 .

Then the animal is injected intravenously with a solution of MNS stabilized by HSA. The entered dose of MNP in terms of iron concentration is 5 mg / kg. MRI scans were performed on a Bruker Clinscan 7T scanner. Then, in T2 * weighted mode with the following parameters: SWI mode: TE / TR = 19/50 ms, slice thickness 0.5 mm, FOV = 30 mm, resolution 256/176 5 minutes after the administration of the MNP preparation.

Obtained during the experiments of the NP in examples 1-5 and have a size of less than 10 nm (Fig. 1)

With an increase in boiling time, a gradual increase in the size of nanoparticles from 7.5 nm to 50 nm occurs (Fig. 2-5).

In addition, with the introduction of MNPs to animals bearing experimental tumors and MRI scans, several types of tumors can be visualized, in particular C6 rat brain glioblastoma, RS-1 liver mucosa, mouse 4T1 mammary adenocarcinoma (Fig. 6-8).

Claims (11)

1. A method of obtaining a preparation for MRI diagnosis of tumor diseases, comprising preparing a solution of iron (III) acetylacetonate in benzyl alcohol with a concentration of 75-200 g / l, followed by heating in an inert gas stream to the boiling point of benzyl alcohol for 4-8 hours, and boiling the solution from 30 minutes to 4 hours to obtain a suspension, then the suspension was cooled, washed with a polar organic solvent to obtain iron oxide nanoparticles of Fe ₃ O _4, which is then coated with human serum albumin and / or ychim serum albumin, and the resulting coating is stabilized intermolecular crosslinking by glutaraldehyde. 2. The method according to p. 1, characterized in that for coating the nanoparticles with human serum albumin and / or bovine serum albumin, the nanoparticles are dissolved in water with a pH of 10-11 to obtain a concentration of 2-8 mg / ml, BSA and / or HSA in the form of an aqueous solution with a concentration of the mentioned BSA and / or HSA 4-16 mg / ml in a ratio of 1: 1 by volume, followed by dialysis of the resulting solution. 3. The method according to p. 1, characterized in that after stabilization of the coating by intermolecular crosslinking, the by-products of the reaction are purified by ultrafiltration and sterilization by filtration. 4. The method according to p. 3, characterized in that the ultrafiltration is carried out on filters with a pore diameter of 100-300 kDa. 5. The method according to p. 1, characterized in that the heating is carried out at a constant speed. 6. The method according to p. 1, characterized in that as a polar organic solvent using water-soluble aliphatic monohydric alcohols, acetones, ketones, nitriles. 7. The method according to p. 6, characterized in that ethanol, methyl alcohol propyl alcohol are used as water-soluble aliphatic monohydric alcohol, acetone, butanone-2 are used as ketones, acetonitrile is used as nitriles. 8. The method according to p. 1, characterized in that the washing is carried out until the disappearance of traces of benzyl alcohol. 9. The method according to p. 8, characterized in that the washing is carried out in volumes equal to at least the volume of the suspension, at least three times. 10. The method according to p. 1, characterized in that the washing is carried out by sedimentation by centrifugation. 11. The method according to p. 1, characterized in that the resulting preparation is freeze-dried.

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