Dosage form for inhibiting bacterial infections and method for organizing same

Abstract

The claimed technical solution relates to the field of pharmaceutics and to dosage forms for expanding the existing range of medicaments based on a gramicidin S-containing pharmaceutical substance for use in inhibiting bacterial infections in a local inflammation site in the human oral cavity and in increasing the effectiveness of treatment of inflammatory infectious diseases of the oral cavity, specifically alveolitis, pharyngitis, periodontal disease, gingivitis and stomatitis, and further relates to methods for organizing said dosage forms. The present dosage form for inhibiting bacterial infections, which is based on a pharmaceutical substance containing gramicidin S, chitosan and hydroxypropyl-β-cyclodextrin, additionally contains a film-forming agent in the form of collagen, and a plasticizer, wherein the dosage form is produced in the form of a biodegradable buccal film having a thickness of 33-37 μm, an aqueous-extract pH of 6.8-7.2 and a T50 index for gramicidin S in relation to human erythrocytes of 24-26 mkg / ml and is capable of prolonged release of gramicidin S. The components of the biodegradable buccal film are present in the following proportions: 2-4 wt% gramicidin S; 30-24 wt% chitosan; 10-15 wt% collagen; 28-32 wt% hydroxypropyl-β-cyclodextrin; 30-25 wt% plasticizer. The claims include 2 independent claims and 8 dependent claims.

Description

[0001] Dosage form for suppression of bacterial infections, method of its organization

[0002] The claimed technical solution relates to the field of pharmaceuticals, to dosage forms that expand the arsenal of drugs based on a pharmaceutical substance containing gramicidin C, which can be used to suppress bacterial infections in the focus of local inflammation of the human oral cavity, to increase the effectiveness of therapy for infectious and inflammatory diseases of the oral cavity, namely alveolitis; pharyngitis; periodontosis; gingivitis; stomatitis and methods for organizing them.

[0003] Terms used:

[0004] Gramicidin C - Gramicidin C hydrochloride or dihydrochloride, an original domestic antibiotic, first obtained by I.F. Gauze and M.G. Brazhnikova in 1942. Gramicidin C hydrochloride is colorless needle-shaped crystals. Gramicidin C exhibits increased antimicrobial activity, is intended exclusively for local use, and has no analogues for systemic use. The drug has a pronounced bactericidal effect on pathogenic gram-positive and gram-negative bacteria, which in most cases are either directly the cause of diseases of the throat and oral cavity, or are added secondarily to the initial viral infection - Streptococcus spp., Staphylococcus spp., Streptococcus pneumoniae, Neisseria, anaerobes, etc. (Khokhlov A.S., Shemyakin M.M. Chemistry of antibiotic substances. - 1949. -455).

[0005] Organize (French organiser, from Latin organum organ) - To connect different objects or forces of nature to achieve a specific goal; to arrange, to revive (Chudinov's Dictionary of Foreign Words of the Russian Language / ZURL: URL: http: / / dic.academic.ru / / ).

[0006] Organization (French: Organisation, from late Latin: organize - I impart a harmonious appearance, I arrange) is a set of processes or actions leading to the formation and improvement of relationships between parts of a whole (Large Encyclopedic Dictionary / ZURL: http: / / dic.academic.ru / / ).

[0007] Dosage form - a state of a medicinal product that corresponds to the methods of its administration and use and ensures the achievement of the required therapeutic effect (State Pharmacopoeia of the Russian Federation [Electronic resource] - 14th edition. - Volume I. - Moscow, 2018. - URL: http: / / www.femb.ru / feml).

[0008] A polymer is a substance with a high molecular weight, the bond between the atoms of which is presented in the form of a linear or branched chain, as well as in the form of a three-dimensional structure. Within a polymer, a repeating structural element, a monomer, can often be distinguished. Depending on their origin, polymers can be natural (proteins, nucleic acids, polysaccharides, rubber and other organic substances) and synthetic (A brief electronic reference book on the main oil and gas terms with a cross-reference system - Moscow: Gubkin Russian State University of Oil and Gas. M.A. Mokhov, L.V. Igrevskiy, E.S. Novik. 2004. / URL: https: / / neft.academic.ru / )

[0009] Films are a dosage form consisting of single or multilayer thin plates of a size suitable for use, containing one or more active substances and auxiliary substances, including film-forming substances (State Pharmacopoeia of the Russian Federation [Electronic resource] - 14th edition. - Volume I. - Moscow, 2018. - URL: http: / / www.femb.ru / feml).

[0010] Adhesion - (from Latin Adhaesio - sticking), the emergence of a bond between the surface layers of two dissimilar (solid or liquid) bodies (phases) brought into contact. It is the result of intermolecular interaction, ionic or metallic bonds. (Physical Encyclopedic Dictionary - Moscow: Soviet Encyclopedia. Chief Editor A. M. Prokhorov. 1983 / URL: https: / / dic.academic.ru / dic.nsf / enc_physics / )

[0011] Technical solutions are known in the form of lozenges, in which the active pharmaceutical substance Gramicidin C is in a solid state of aggregation. (Patent RU 2213558 C1, IPC A61 K 9 / 20, A61 K 38 / 08, priority 09 / 30 / 2002, published 10 / 10 / 2003; Patent RU 2160088 C1, IPC A61K 7 / 16, A61 K 9 / 06, priority 02.11.1999, published 10.12.2000; Patent RU 2590980 C2, IPC A61 R 31 / 00, A61 K 31 / 155, A61 K 31 / 7004, A61 K 31 / 194, A61 K 31 / 375, A61 K 33 / 30, priority 14 / 24 / 2009, published 10.07.2016).

[0012] The disadvantages of known technical solutions include the low bioavailability of Gramicidin C in the local inflammation focus. When entering the oral cavity, solid particles of Gramicidin C are not able to firmly bind to the surface of the infection focus and are washed out from this surface, which reduces the therapeutic effect of known antibacterial compositions. In addition, they contain auxiliary substances that can cause side effects, such as allergic reactions (lactose, sucrose).

[0013] There are known technical solutions in which the active pharmaceutical substance Gramicidin C is in a liquid aggregate state, which is an alcohol solution from light yellow to yellow with a characteristic odor, containing 2% Gramicidin C hydrochloride, which has a specific taste and odor. (Gramicidin C - official instructions for use, URL: https: / / medi.ru / instrukciya / gramitsidin-s_12289 / ).

[0014] The disadvantage of the known technical solution is that, as a result of diluting a 2% alcohol solution of Gramicidin C during the preparation of the drug, opalescence of the solution is observed with subsequent formation of a precipitate, which leads to a decrease in therapeutic properties, since Gramicidin C takes the form of solid particles. Under industrial conditions, it is unprofitable and impossible to produce an aqueous solution of Gramicidin C based on a 2% alcohol solution due to the short duration of preservation of the aggregate stability of Gramicidin C particles in aqueous solutions, which does not exceed 3 days. The preparation of a drug based on a 2% alcohol solution at home is also impossible due to the lack of the ability to accurately dose the components. The listed facts exclude the possibility of retail sale of a 2% alcohol solution of Gramicidin C. It is also known that the method of using the resulting aqueous solution of Gramicidin C (rinsing) is not effective enough, sincethe active substance is not fully distributed in the infection site, but partially goes beyond it, which does not allow for precise dosing of the active substance. In this regard, it is considered more effective to create a liquid dosage form containing Gramicidin C in the form of a spray, which allows for a higher concentration of the active substance in the infection site and ensures its precise dosing.

[0015] Technical solutions are known that describe an antibacterial composition based on a pharmaceutical substance including Gramicidin C and ethanol for the treatment of infectious and inflammatory diseases of local application, a method for its production and use (Patent RU 2604576 C1, IPC A61 K 31 / 216, A61 K 31 / 44, A61 K 38 / 12, A61 K 9 / 12, A61 K 2121 / 00 priority 01.12.2015, published 10.12.2016, Patent RU 2604575 C1, IPC A61K 31 / 216, A61 K 31 / 44, A61 K 38 / 12, A61 K 9 / 12, A61 K 2121 / 00 priority 01.12.2015, published 12 / 10 / 2016, Patent RU 26267423 C1, IPC A61 K 38 / 12, A61 K 31 / 44 A61 K 9 / 12, A61 P 31 / 00 priority 06 / 29 / 2016, published 08 / 08 / 2017, Patent RU 2749902 C2, IPC A61 K 38 / 12, A61 K 31 / 137, A61 R 31 / 00, A61 R 31 / 04, A61 R 11 / 00, A61 R 11 / 04 priority 10 / 31 / 2018, published 06 / 16 / 2021 ).

[0016] The composition contains Gramicidin C, oxybuprocaine hydrochloride and cetylpyridinium chloride, ethanol 96%, as auxiliary substances - sucralose, glycerol, citric acid monohydrate, sodium citrate, polysorbate 80, methylparaben, propylparaben, mint flavoring, purified water.

[0017] A technical solution is known that describes an antibacterial composition for delivering Gramicidin C to the site of local inflammation, made in the form of a medicinal solution, with the possibility of maintaining it in the state of a medicinal solution until use, a method for its production and use (Patent RU 2659418 C1, A61 K 38 / 12, A61 K 47 / 10, A61 K 47 / 14, A61 K 9 / 12, A61 P 31 / 04 priority 08 / 24 / 2017, published 07 / 02 / 2018).

[0018] The disadvantage of the known technical solutions is the high probability of allergic reactions and side effects when used. The occurrence of these effects may be due to the presence of such substances as oxybuprocaine hydrochloride, cetylpyridinium chloride, citric acid monohydrate, sodium citrate, methylparaben and propylparaben in the composition of the known technical solutions. It is known that oxybuprocaine hydrochloride can cause temporary burning of the mucous membranes with prolonged use. Also, in the presence of increased sensitivity of the body, the component can cause adverse reactions from the cardiovascular system, anaphylactic shock, temporary or short-term loss of consciousness. Cetylpyridinium chloride causes local irritation with prolonged use. In some cases, it contributes to the development of gastrointestinal disorders, noted mainly when exceeding the recommended doses.Citric acid monohydrate and sodium citrate also cause local irritation of the mucous membrane. Methylparaben and propylparaben, due to their surface-active properties, are able to penetrate deep into the tissues of the body, accumulate (especially in case of impaired renal function) and initiate the development of breast cancer, male fertility. Additionally, parabens can cause damage to the mucous membrane and cause allergic reactions in the form of skin rash and itching. The negative effect of the use of these substances is exacerbated by damage to the tissues of the oral cavity and throat. A technical solution is known that describes the preparation of lyophilisates of hydroxypropylcyclodextrin complexes using co-solvents (Patent US 5120720, IPC A61K 9 / 18 priority 20.01.1990, published 09.06.1992). The known technical solution discloses a method for increasing the solubility of substances, incl. gramicidin C, in water through the formation of an inclusion complex.

[0019] The disadvantage of the known technical solution is that the resulting complexes have low adhesion to the mucosal surface and, as a consequence, low therapeutic activity.

[0020] Overcoming this drawback is disclosed in the known technical solution selected as a prototype - Pharmaceutical substance for the formation of medicinal products based on Gramicidin C and the method for its production (variants), pharmaceutical substance for the formation of prolonged-release medicinal products based on Gramicidin C and the method for its production (variants) (Patent RU 2733715 C2, IPC A61K 38 / 12, A61K 31 / 722, A61K 31 / 724, A61K 9 / 20, A61K 9 / 10, B01F 3 / 08 priority 01 / 18 / 2019, published 10 / 06 / 2020).

[0021] The implementation of the technical solution according to the prototype allows for a prolonged action of gramicidin C due to the formation of a complex with β-cyclodextrin or its analogues and chitosan, which has increased adhesion to the tissues of the human body and, as a consequence, longer retention of gramicidin C at the site of infection.

[0022] The disadvantage of the known technical solution is the fact that the amount of pharmaceutical substance according to the prototype, necessary to ensure a therapeutic effect, is extremely small, which makes it impossible to directly use the pharmaceutical substance for treatment and causes the need to form a dosage form that will allow the manipulations necessary for treatment to be performed.

[0023] A technical solution is known - prolonged medicinal forms of local action, which have an activating effect on the immune system (Patent RU 2185807 C1, IPC A61 K 6 / 00, A61K 9 / 70, priority 27.07.2001, published 27.02.2002). The known technical solution is a two-layer film, which is intended for the treatment of diseases of the oral cavity. The disadvantage of the known technical solution is the presence of a hydrophobic layer in the composition, which practically does not dissolve in the oral cavity environment and must be removed from the surface of the infection site by manipulation.

[0024] In this regard, the authors were tasked with developing a new dosage form that would expand the range of products based on a pharmaceutical substance containing gramicidin C, which would have a local effect on the site of infection, which would have the ability to release gramicidin C in a prolonged manner, and which, as the active substance is released, would be completely dissolved in the oral cavity, and which would also have a reduced level of hemolysis of gramicidin C in relation to human erythrocytes, and a method for organizing it.

[0025] The problem is solved in that the dosage form for suppressing bacterial infections, made on the basis of a pharmaceutical substance including gramicidin C, chitosan and hydroxypropyl-P-cyclodextrin, additionally contains a film former in the form of collagen and a plasticizer, while it is made in the form of a biodegradable cheek film with a thickness of 33-37 μm, with a pH of the aqueous extract of 6.8-7.2, a Tso value of gramicidin C in relation to human erythrocytes of 24-26 μg / ml, and has the ability to prolonged release gramicidin C.

[0026] The ratio of components in the biodegradable cheek film in May is:

[0027] Gramicidin E 2 - 4

[0028] Chitosan 30 - 24

[0029] Collagen 10 - 15

[0030] Hydroxypropyl-P-cyclodextrin 28 - 32

[0031] Plasticizer 30 - 25

[0032] Gramicidin C may be gramicidin C base, gramicidin C dihydrochloride. The plasticizer may be glycerin, propylene glycol, polyvinyl alcohol. The biodegradable cheek film is a platelet that may be in a solid or gelled state.

[0033] In a method for organizing a dosage form for suppressing bacterial infections, including the use of a pharmaceutical substance containing gramicidin C, chitosan and hydroxypropyl-p-cyclodextrin, a film former in the form of collagen and a plasticizer are additionally used; the film formation process is carried out, maintaining the moisture removal rate at a level of 0.5% / min, with the production of a biodegradable cheek film with a thickness of 33-37 μm, with a pH of the aqueous extract of 6.8-7.2, a Tso indicator of gramicidin C in relation to human erythrocytes of 24-26 μg / ml, having the ability to delay the release of gramicidin C.

[0034] In this case, the ratio of components in the biodegradable cheek film is selected in May. %:

[0035] Gramicidin E 2 - 4

[0036] Chitosan 30 - 24

[0037] Collagen 10 - 15

[0038] Hydroxypropyl-p-cyclodextrin 28 - 32

[0039] Plasticizer 30 - 25

[0040] Gramicidin C may be gramicidin C base, gramicidin C dihydrochloride. The plasticizer may be glycerin, propylene glycol, polyvinyl alcohol. The biodegradable cheek film is a platelet that may be in a solid or gelled state.

[0041] The technical effect of the claimed technical solution consists in expanding the arsenal of means for this purpose, reducing the level of hemolysis of gramicidin C in relation to human erythrocytes.

[0042] The essence of technical solutions is explained as follows.

[0043] Gramicidin C has been used in clinical practice for over 50 years, during which it has demonstrated effectiveness against various bacteria. At the same time, the main form of release of gramicidin C is lozenges, the effectiveness of which in case of local therapy is low due to the uniform distribution of the active substance over the entire surface of the oral mucosa, including uninfected areas, which can cause an irritating effect. In the case of using sprays, the sprayed solution does not contain components that will promote its adhesion to the surface of the mucosa, which will lead to rapid washing out of the active substance from its surface.In this regard, it is becoming relevant to develop a product containing gramicidin C in the form of a film consisting of mucoadhesive and biodegradable polymers, which will allow for longer retention of the product on the surface of the infection site when applied locally and will reduce the level of side effects due to the decomposition of the film base over time.

[0044] When organizing a dosage form for suppressing bacterial infections, the following sequence of technological operations is used:

[0045] - Dissolution of acetic acid in purified water to obtain a 1% solution of acetic acid

[0046] - Dissolution of chitosan and hydroxypropyl-p-cyclodextrin in a 1% solution of acetic acid to obtain a polymer solution.

[0047] - Dissolution of gramicidin C in a polymer solution to obtain a solution of the active substance.

[0048] -Mixing a solution of the active substance, a 0.1% solution of collagen in 0.1 M acetic acid and a plasticizer to obtain a solution of biodegradable cheek film.

[0049] -Homogenization of the biodegradable cheek film solution at 10,000 rpm.

[0050] - Formation of biodegradable cheek films with removal of acetic acid and purified water

[0051] During the film formation, the moisture removal rate is maintained at 0.5% / min. In this case, the thickness of the biodegradable cheek film is 33-37 μm, with a pH of the aqueous extract of 6.8-7.2, a Tso value of gramicidin C in relation to human erythrocytes of 24-26 μg / ml, with the ability to delay the release of gramicidin C.

[0052] The ratio of components in biodegradable cheek film in May % is chosen:

[0053] Gramicidin E 2 - 4

[0054] Chitosan 30 - 24

[0055] Collagen 10 - 15

[0056] Hydroxypropyl-₽-cyclodextrin 28 - 32

[0057] Plasticizer 30 - 25

[0058] Example 1 To confirm the possibility of obtaining a dosage form for suppressing bacterial infections, the authors demonstrate the possibility of implementing the claimed method in relation to the following compositions

[0059] Continuation of example 1 Continuation of example 1

[0060] The organization of samples of the dosage form for the suppression of bacterial infections is carried out using a sequence of technological operations:

[0061] - Dissolve acetic acid in purified water to obtain a 1% solution of acetic acid

[0062] - Chitosan and hydroxypropyl-3-cyclodextrin are dissolved in a 1% acetic acid solution to obtain a polymer solution.

[0063] - Dissolve gramicidin C in a polymer solution to obtain a solution of the active substance.

[0064] - Mix the active substance solution and 0.1% collagen solution in 0.1 M acetic acid to obtain a solution of biodegradable cheek film.

[0065] -Mix the film and plasticizer solution to obtain a biodegradable cheek film solution.

[0066] -Homogenize the biodegradable cheek film solution at 10,000 rpm.

[0067] Biodegradable cheek films are formed by removing acetic acid and purified water, maintaining the moisture removal rate at 0.5% / min. to obtain a dosage form for suppressing bacterial infections. In this case, the thickness of the biodegradable cheek film is 33-37 μm, with a pH of the aqueous extract of 6.8-7.2, a Tso value of gramicidin C in relation to human erythrocytes of 24-26 μg / ml, with the ability to delay the release of gramicidin C. When organizing a dosage form for suppressing bacterial infections, it is necessary to maintain the ratio of components, which will ensure the stability of the dosage form (options 2.5, 8, 11, 14).

[0068] If the content of gramicidin C deviates (options 1, 3), the therapeutic effectiveness of the dosage form for suppressing bacterial infections is impaired, since the required amount of the active substance is not provided or there is an excess of it, which can cause a side effect.

[0069] If the percentage content of film-forming components - chitosan and collagen (options 4, 6, 7, 9) deviates, the dosage form for suppressing bacterial infections will differ in thickness from the target value of 33-37 microns.

[0070] The content of hydroxypropyl-|3-cyclodextrin causes the dissolution of gramicidin C in 1% acetic acid solution when obtaining a polymer solution. In case of a deficiency of hydroxypropyl-β-cyclodextrin, gramicidin C will not dissolve in 1% acetic acid solution (option 10), which will lead to deterioration of the dosage uniformity of the drug form for suppressing bacterial infections. In case of an excess of hydroxypropyl-β-cyclodextrin (option 12), this substance will have a drying effect on the film, which will lead to deterioration of elasticity.

[0071] Elasticity, in turn, can be regulated by the content of plasticizer. If there is a deficiency of this component (options 13, 16, 19), an inelastic film with a low tensile strength is formed. In case of excess plasticizer (options 15, 18, 21), problems with film formation arise, since moisture removal is insufficient, and the solution of biodegradable cheek film does not pass into the medicinal form for suppressing bacterial infections.

[0072] Example 2

[0073] The adhesive properties of the dosage form for suppressing bacterial infections were determined using the rheological method (Kirzhanova, E. A. Methods of mucoadhesion analysis: from fundamental research to practical application in the development of dosage forms / E. A. Kirzhanova, V. V. Khutoryansky, N. G. Balabushevich, A. V. Kharenko, N. B. Demina / / Development and registration of drugs. - 2014. - No. 8 (3). - P. 66) on a Smart L rotational viscometer. Mucin was dissolved in 10 ml of phosphate buffered saline to obtain a 5% solution.

[0074] A 5% mucin solution containing 10.0 mg of chitosan was prepared separately.

[0075] A 5% mucin solution containing glycerol weighing 10.5 mg was prepared separately.

[0076] A 5% mucin solution containing 10.0 mg of collagen was prepared separately.

[0077] A 5% mucin solution containing 2.0 mg of hydroxypropyl-β-cyclodextrin was prepared separately. A 5% mucin solution containing a complex of gramicidin C and hydroxypropyl-β-cyclodextrin was prepared separately. A 5% mucin solution containing all components of the bacterial infection suppression dosage form was prepared separately. The viscosity of the resulting solutions was measured at 37 °C and a spindle speed of L4 of 30 rpm, and then the mucoadhesion viscosity for the bacterial infection suppression dosage form was determined using equation 1:

[0078] G|mucoadhesions = G|complex - G|mucin > G|GP-R-CD > Г|ГС (1 ) for film - using equation 2:

[0079] G|mucoadhesions =G|film - G|mucin - G|GP-₽-CD - G|GS - G|chitosan- G|collagen - G|glycerol (2) where n complex is a complex of gramicidin C and hydroxypropyl-β-cyclodextrin; Pmucin is the viscosity of a 5% mucin solution; g|gp-₽-CD is the viscosity of a hydroxypropyl-β-cyclodextrin solution; Pgs is the viscosity of a gramicidin C solution; Pchitosan is the viscosity of a chitosan solution; G]collagen is the viscosity of a collagen solution; Pglycerol is the viscosity of a glycerol solution; G]film - the viscosity of a film solution;

[0080] Pmucoadhesion is the viscosity of mucoadhesion, which is caused by the mucoadhesive interaction of mucin and the test sample, or the strength of mucoadhesion.

[0081] As a result, the following data were obtained:

[0082]

[0083] From the obtained results, it is possible to observe a general increase in the adhesion of the film to the component of the mucous membrane, which will allow the use of gramicidin C preparations in a more targeted manner, creating a depot of the active substance.

[0084] Example 3

[0085] The release of gramicidin C was studied in a medium simulating the oral cavity environment, prepared in accordance with the requirements of the State Pharmacopoeia of the Russian Federation, XIV edition:

[0086] • Phosphate buffered saline pH 7.4

[0087] A conical flask was filled with 700 mg of the bacterial suppression dosage form and 80 ml of the release media were added. The tightly closed flask was placed in a dry-air laboratory thermostat and the release process was carried out at 37 °C and stirring at 100 rpm for 12 h. At specified time intervals, 1.5 ml of the release medium were sampled. The sampled volume was compensated with a fresh solution with a specified pH value and the process was continued. The sample was dried under vacuum at 40 °C and extracted with ethyl alcohol using ultrasound, determining the amount of released gramicidin C using the HPLC method.

[0088] The percentage of released gramicidin C (R) was determined using formula 3:

[0089] R (%) = D t / Dmax*100, where (3)

[0090] Dt is the concentration of gramicidin C in the solution at the moment in time (mg / ml);

[0091] Dmax is the maximum possible amount of gramicidin C in solution (mg / ml).

[0092] The result is long-lasting.

[0093] Example 4

[0094] To determine the pH of an aqueous solution of the dosage form for suppressing bacterial infections, 0.5 g of ground films were placed in a 50 ml conical flask, 25 ml of water were added, heated in a water bath until dissolved, cooled to room temperature and filtered through a blue ribbon paper filter, followed by measurement of the pH of the filtrate.

[0095] As a result, it was found that the pH of the solution of the dosage form for suppressing bacterial infections has a value of 6.8-7.2, which is as close as possible to the pH value of saliva. Thus, the film will have a minimal irritating effect on the mucous membrane due to the creation of an optimal environment at the site of release of gramicidin C.

[0096] Example 5

[0097] Further evidence of the reduction in the level of side effects is the reduction in the level of hemolysis of gramicidin C in relation to human erythrocytes.

[0098] The study was performed according to generally accepted procedures (Di Martino, A. Folic acid-chitosan-alginate nanocomplexes for multiple delivery of chemotherapeutic agents / A. Di Martino, ME Trusova, PS Postnikov, V. Sedlarik / / Journal of Drug Delivery Science and Technology. - 2018. - No. 47. - Pp. 67-76) using a freshly collected blood sample in the amount of 3 ml. At the first stage, the blood was washed by adding the collected sample to 14 ml of phosphate buffer with pH 5.7 and mixing by shaking on an orbital shaker, after which the resulting suspension was centrifuged for 10 minutes at 500 g and a temperature of 4 °C, using a multifunctional centrifuge. The supernatant was removed and the above sequence of operations was repeated 4 times, after which a concentrated blood solution with a concentration of 2% was prepared by collecting 0.2 ml of blood from the bottom of the vial and adding it to 9.8 ml of phosphate buffer with pH 5.7.Separately, the test aqueous solutions of gramicidin C and the dosage form for suppressing bacterial infections were prepared by serial dilutions, obtaining the concentration values ​​of the active substance in the solution of 500 μg / ml, 250 μg / ml, 125 μg / ml, 62.5 μg / ml, 31.25 μg / ml, 15.625 μg / ml. To obtain an aqueous solution of gramicidin C, the initial suspension with a concentration of 500 μg / ml was treated with ultrasound. The concentration of gramicidin C dihydrochloride in the test solutions was confirmed by the results of quantitative analysis by HPLC. To determine the level of hemolysis, 50 μl of a 2% blood solution were added to 100 μl of the test solutions at different concentrations and incubated for 30 minutes at 37 °C in a thermostat, after which the samples were centrifuged at 2500 g for 6 minutes, 0.1 ml of the supernatant was collected and diluted with a phosphate buffer solution with pH 5.7 for analysis.The obtained samples were analyzed using a UV spectrophotometer, assessing the absorption of the samples at a wavelength of 541 nm and comparing the signal with the lysis level. As a positive control (lysis level), a solution obtained by the following method was used: 50 μl of 2% blood solution was added to 100 μl of distilled water, subjecting the resulting sample to three freeze-thaw cycles at -20 °C.

[0099] Where a are the values ​​for the dosage form for suppressing bacterial infections, b are the values ​​for gramicidin C.

[0100] From the data of Example 6, it is evident that Tso for human erythrocytes in the presence of gramicidin C in the composition of the dosage form for suppressing bacterial infections increases to a level of 24-26 μg / ml from 11 μg / ml, which significantly reduces the risk of hemolysis when using the dosage form for suppressing bacterial infections.

[0101] The advantages of the claimed technical solution are as follows:

[0102] - Reduced level of gramicidin-induced hemolysis of human erythrocytes;

[0103] - Possibility of local application of gramicidin C to the site of local infection of the oral cavity, increased bioavailability; - No need to extract the dosage form, since it is completely decomposed in the oral cavity during treatment.

Claims

Invention formula 1. A dosage form for suppressing bacterial infections, made on the basis of a pharmaceutical substance including gramicidin C, chitosan and hydroxypropyl-®-cyclodextrin, characterized in that it additionally contains a film-forming agent in the form of collagen and a plasticizer, while it is made in the form of a biodegradable cheek film with a thickness of 33-37 μm, with a pH of the aqueous extract of 6.8-7.2, a Tso value of gramicidin C in relation to human erythrocytes of 24-26 μg / ml, and having the ability to prolonged release gramicidin C, while the ratio of components in the biodegradable cheek film in May is Gramicidin E 2 - 4 Chitosan 30 - 24 Collagen 10 - 15 Hydroxypropyl-p-cyclodextrin 28 - 32 Plasticizer 30 - 25 1. The dosage form according to paragraph 1, characterized in that the dosage form according to paragraph 2, characterized in that gramicidin C is a gramicidin C base, gramicidin C dihydrochloride.

2. The dosage form according to paragraph 2, characterized in that the plasticizer is glycerin, propylene glycol, or polyvinyl alcohol.

3. The dosage form according to paragraph 1, characterized in that the biodegradable buccal film is in the form of thin plates that are in a solid or gelled state.

4. A method for preparing a dosage form for suppressing bacterial infections, comprising using a pharmaceutical substance containing gramicidin C, chitosan and hydroxypropyl-[3-cyclodextrin, characterized in that a film former in the form of collagen and a plasticizer are additionally used; the film formation process is carried out by maintaining the moisture removal rate at 0.5% / min, resulting in a biodegradable cheek film with a thickness of 33-37 μm, with an aqueous extract pH of 6.8-7.2, a Tso value of gramicidin C in relation to human erythrocytes of 24-26 μg / ml and the ability to prolonged release of gramicidin C, while selecting the ratio of components in the biodegradable cheek film in May. % Gramicidin E 2 - 4 Chitosan 30 - 24 Collagen 10 - 15 Hydroxypropyl-P-cyclodextrin 28 - 32 Plasticizer 30 - 25 5. The method according to paragraph 4, characterized in that the method according to paragraph 5, characterized in that gramicidin C is selected as a gramicidin C base, gramicidin C dihydrochloride.

6. The method according to paragraph 4, characterized in that glycerin, propylene glycol, or polyvinyl alcohol are selected as the plasticizer.

7. The method according to paragraph 4, characterized in that the biodegradable cheek film is made in the form of thin plates that are in a solid or gelled state.

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