RU2462254C2 - Silver nanocomposite of sulphated arabinogalactan exhibiting antimicrobial and antithrombotic activity and method for preparing it - Google Patents

Abstract

FIELD: medicine, pharmaceutics.

SUBSTANCE: group of inventions refers to medicine. A method for preparing a silver nanocomposite of semi-synthetic heparinoid - sulphated arabinogalactan (AGSO₃H) in the form of the silver-containing system (AGSO₃H +Ag°) of zero-valent metallic silver nanoclusters of particle size 10-15 nm (X-ray diffraction analysis) stabilised by sulphated arabinogalactan (AGSO₃H) by the reaction of sulphated arabinogalactan in an aqueous medium for 30-60 minutes at temperature 20°C and silver oxide (Ag₂O) accompanied by the product recovery. The silver content evaluated by atomic absorption analysis makes 8%.

EFFECT: group of inventions provides maintaining the structural organisation of silver nanocomposite and sulphated arabinogalactan (nanostructuredness of the biopolymer macromolecule system - particle oversize sinter size makes 200-600 nm), as well as water-solubility, biocompatibility, receptor-mediated transmembrane and immunomodulatory properties in relation or a living cell, antithrombotic activity, prolonged biological action, non-toxicity, synergism of antibacterial activity of silver.

2 cl, 3 dwg, 3 ex

Description

The invention relates to the pharmaceutical industry, medicine, pharmacology, and for the production of silver nanocomposite with antimicrobial and antithrombotic activity, based on semisynthetic heparinoid - sulfated arabinogalactan (AGSO ₃ H). The developed compound can be used in practical medicine as a highly effective antibacterial agent and a wide range of wound coverings with low toxicity, and in combination with antithrombotic action can be used to create special coatings for implants (cardiovascular surgery, neurosurgery, etc. .).

In the literature, methods are known for producing nanosized silver particles stabilized by an arabinogalactan matrix (Patents RU, 2260500 and 2278669).

Closest to the proposed method for producing a silver nanocomposite and sulfated arabinogalactan is a method for producing an antimicrobial agent (RU 2278669), where a polysaccharide - arabinogalactan is used as a stabilizing high molecular weight matrix of ultrafine silver-containing systems. The synthesis of nullovalent metallic silver nanoclusters with a particle size of 10-30 nm, stabilized with arabinogalactan, is carried out by its interaction with silver nitrate (AgNO ₃ ) in an aqueous medium for 30-90 minutes at a temperature of 20 ° C, in a highly alkaline environment (30% aqueous solution of ammonia or sodium hydroxide) at a pH of 10-11, with incubation of solutions at a temperature of 20-90 ° C for 5-60 minutes, with further filtration and precipitation in rectified ethyl alcohol.

This method has several disadvantages.

1. The method of producing ultrafine silver-containing systems stabilized with arabinogalactan is long, laborious, requires the use of high temperatures and a very strong alkaline environment.

2. The stabilizing matrix - arabinogalactan can effectively stabilize metallic silver particles, making them water-soluble, but they do not acquire new biologically active properties that can be provided by sulfated arabinogalactan, which, in turn, has the property of heparinoid: anticoagulant, antithrombotic, etc. .d. activities that are described in patent RU 2319707.

The aim of our invention is the synthesis of silver nanocomposite and sulfated arabinogalactan, which is a nanovalent silver nanoclusters with a particle size of 10-15 nm, stabilized with sulfated arabinogalactan, which has not only antimicrobial activity, but also antithrombotic activity. A distinctive feature of the compound developed by us is the synergism of the antibacterial activity of silver and the properties of the semisynthetic heparinoid - sulfated arabinogalactan (water solubility, biocompatibility, receptor-mediated transmembrane properties with respect to a living cell, immunomodulating properties, antithrombotic activity, prolonged biological activity, nano-thrombolecule, nontromolecularity, non-toxicity) (size of large particle agglomerates composition yaet about 200-600 nm), allowing to obtain compounds with high bioavailability, which is manifested in high efficiency with a minimum of side effects.

Describes a method for producing ultrafine silver-containing systems (AGSO ₃ H + Ag ⁰ ), which are nanoclusters of nullovalent metallic silver with a particle size of 10-15 nm, stabilized with sulfated arabinogalactan (AGSO ₃ H), by its interaction with silver oxide (Ag ₂ O ) in an aqueous medium for 60 minutes at a temperature of 20 ° C with further filtering and planting in ethyl rectified alcohol.

The technical result of the present invention is to obtain a stable water-soluble compound in solid powder form containing nanosized silver particles, preserving the pharmacological activity characteristic of heparinoids, in particular sulfated arabinogalactan, and at the same time possessing antimicrobial activity.

This goal is achieved by the fact that silver oxide (Ag ₂ O) was added to an aqueous solution of sulfated arabinogalactan to obtain null-valent metallic silver nanoclusters with a particle size of 10-15 nm, the reaction was carried out for 60 minutes at a temperature of 20 ° C with further filtering and planting in rectified ethyl alcohol.

The silver content in the obtained sample, determined by atomic absorption analysis, is 8%.

According to electron microscopy, silver in the ultrafine system stabilized by sulfated arabinogalactan is in a zero-valence state and its particle size is 10-15 nm.

The proposed method is characterized by the following advantages:

- obtaining a nanocomposite of silver and sulfated arabinogalactan is simple in technical design and economical, since it does not require the use of high temperature and a very strong alkaline environment, as well as a large investment of working time;

- the resulting nanocomposite based on sulfated arabinogalactan preserves the structural organization, water solubility and a wide range of pharmacological activity of heparinoid.

Fig. 1 shows transmission electron microscopy of a silver nanocomposite and sulfated arabinogalactan.

Figure 2 presents a table showing the antimicrobial activity of silver nanocomposite and sulfated arabinogalactan.

Fig. 3 is a diagram showing the effect of silver nanocomposite and sulfated arabinogalactan on platelet aggregation.

The following examples illustrate the invention.

Example 1

0.2 g of AGSO ₃ H was dissolved with stirring in 3 ml of distilled water, then 0.15 g of Ag ₂ O was added gradually with stirring. After 60 minutes, the resulting product was filtered off from unreacted Ag ₂ O through a paper filter (blue ribbon) and planted in 20 ml of rectified ethyl alcohol. The yield of the obtained product in the form of a powder of dark brown color was 98%. The quantitative content of silver according to elemental analysis is 8%.

Example 2

The antimicrobial effect of silver nanocomposite and sulfated arabinogalactan was studied by the replication method (Gunar O.V., Kagramanova K.A. // Chem. - Pharmaceutical journal. - 2005. - T.39. - No. 5. - P.53-56 ) recommended for stained substances on test microorganisms obtained from the State collection of pathogenic microorganisms GISK them. L.A. Tarasevich, and strains isolated from patients from the purulent-septic center of the Scientific Center for Internal Medicine of the Siberian Branch of the Russian Academy of Medical Sciences with various surgical pathologies.

In sterile Petri dishes, 1 ml of the corresponding solutions of silver nanocomposite and sulfated arabinogalactan in a concentration of 0.1-3% prepared in phosphate buffer (pH 7.6) was added, 15 ml of molten and cooled MPA or Saburo medium were added. After solidification of the agar, an inoculum of daily cultures of each test strain of microorganisms was applied to the surface of the medium at a concentration of 10 ³ -10 ⁴ CFU / ml in the form of plaques. Nutrient media were used as a control, into which 1 ml of phosphate buffer (pH 7.6) was added instead of the studied solutions. Crops were incubated on MPA at 32-34 ° C for 48 hours, on Saburo at 22-24 ° C for 72 hours. The results of the experiment were taken into account by the presence and growth pattern of test strains of microorganisms on nutrient media. The presence of growth of test microorganisms was indicated by the sign "+", the absence of growth by the sign "-". The results obtained are presented in Table 1 (Figure 2).

The research results showed that the nanocomposite of silver and sulfated arabinogalactan has a pronounced antimicrobial effect. A solution of an ultrafine silver-containing system (AGSO ₃ H + Ag ⁰ ), which is a null-valent metallic silver nanoclusters with a particle size of 10-15 nm, stabilized with sulfated arabinogalactan (AGSO ₃ H), at a concentration of> 3% can inhibit the growth of gram-positive and gram-negative microflora, including fungi of the genus Candida, at a concentration of 1% acts only on gram-negative microflora, at a concentration of <0.5%, the antimicrobial effect is lost.

Example 3

Antithrombotic activity was investigated in vitro by the effect of silver nanocomposite and sulfated arabinogalactan on the platelet aggregation of healthy donor plasma caused by the action of an aggregation inducer, which was used ADP at a concentration of 2.5 μg / ml according to a unified method using a Colar aggregometer and reagents of the company "Technology - Standard" (Manual on the experimental (preclinical) study of new pharmacological substances / Edited by V.P. Fisenko. - M .: ZAO IIA "Remedium", 2000. - 398 p.).

The results of the experiments are presented in Figure 3.

A silver and sulfated arabinogalactan nanocomposite exhibits a therapeutic antiaggregation effect of a concentration of 0.5 mg / ml plasma, which consists in reducing platelet aggregation by an inducer by 50% compared with the control.

Thus, the silver and sulfated arabinogalactan nanocomposite has a pronounced antithrombotic activity and a dose-dependent antimicrobial effect, which can be used to develop highly effective antibacterial agents and wound coatings with a wide spectrum of action, as well as special coatings for implants.

Claims (2)

1. A method of obtaining an ultrafine silver-containing system (AGSO ₃ H + Ag ^o ), characterized by the interaction of silver oxide with sulfated arabinogalactan in an aqueous medium at a temperature of 20 ° C for 30-60 minutes with further isolation of the product, which is a nullvalent metallic silver nanocluster with a size particles of 10-15 nm in a matrix of sulfated arabinogalactan, planting in ethyl alcohol. 2. The ultrafine silver-containing system (AGSO ₃ H + Ag ^o ), obtained according to claim 1, which is a nano-valence metal silver nanocluster with a particle size of 10-15 nm in a sulfated arabinogalactan matrix, having both antimicrobial and antithrombotic activity.

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