Pharmaceutical composition with antibiotic activity

Abstract

The invention relates to pharmaceutics, and more particularly to gastroenterology, and can be used for the prophylaxis or treatment of inflammatory diseases of the gastrointestinal tract. What is described is a pharmaceutical composition having antibiotic activity, characterized in that it contains a therapeutically effective amount of crystalline rifaximin in anhydrous form.

Description

[0001] PHARMACEUTICAL COMPOSITION WITH ANTIBIOTIC ACTIVITY

[0002] Field of technology The invention relates to pharmaceuticals, in particular gastroenterology, and can be used for the prevention or treatment of inflammatory diseases of the gastrointestinal tract.

[0003] Background of the invention

[0004] Rifaximin is a semi-synthetic antibiotic of the rifamycin group, synthesized from natural rifamycin S by bromination to obtain 3-bromorifamycin, followed by the addition of 2-amino-4-methylpyridine, treatment of the reaction mixture with ascorbic acid and crystallization of the final compound of the formula: Rifaximin is practically insoluble in water, when taken orally it remains inside the stomach and intestines, without penetrating their walls, creating a high concentration of the substance inside the esophagus and a very low one in the blood plasma, such properties ensure its high efficiency and safety in the treatment of gastrointestinal tract infections.

[0005] Pure rifaximin is a crystalline powder from dark red to pink, depending on the polymorphic modification. When isolated from organic or water-organic mixtures, it can form various crystal hydrate structures that differ in both water content and crystal shapes and sizes; in addition, an amorphous modification of rifaximin and various methods for obtaining it are known.

[0006] Eurasian patent EA 022324 B1 describes an amorphous powder of rifaximin obtained by spray drying, in which an alcohol solution of rifaximin is fed into a nozzle, dispersed to an aerosol, mixed with hot air and dried to a powder with characteristic amorphous properties. The same patent describes a method for obtaining an amorphous powder by grinding crystalline rifaximin to aggregates of 3-15 microns in size.

[0007] Crystallization of rifaximin from various solvents under certain temperature conditions, stirring intensity and sedimentation rate allows obtaining various polymorphic modifications of the substance.

[0008] Crystallization of rifaximin from aqueous-alcoholic solutions with heating to 45°-65°C and subsequent cooling is described in Russian patent 2397985. The rifaximin obtained in this way has an 8 polymorphic form, which upon drying transforms into the 8 form. The described 8 and 8 polymorphs of rifaximin have different solubility and bioavailability, and therefore can be used to produce various medicinal products that differ in composition and therapeutic action.

[0009] Russian patent 2270200 C2 describes polymorphic modifications of rifaximin A (demonstrating peaks at diffraction angles of 20: 6.6°; 7.4°; 7.9°; 8.8°; 10.5°; 11.1°; 11.8°; 12.9°; 17.6°; 18.5°; 19.7°; 21.0°; 21.4°; 22.1°); | (demonstrating peaks at diffraction angles of 20: 5.4°; 6.4°; 7.0°; 7.8°; 9.0°; 10.4°; 13.1°; 14.4°; 17.1°; 17.9°; 18.3°; 20.9°) and y (demonstrating a mainly amorphous profile and several significant peaks at diffraction angles of 20: 5.0°; 7.1°; 8.4°) and methods for their isolation by crystallization from aqueous-alcoholic solutions.According to the description, rifaximin is dissolved in alcohol at a temperature of 45°-65°C, water is added to the solution, cooled until crystallization begins, then reheated to a temperature of 40°-50°C, held for 6 to 24 hours, cooled to 0°C and the resulting rifaximin crystal hydrates are dried, and depending on the residual moisture, polymorph A with a water content of 2.0%-3.0%, polymorph P with a water content of 5.0%-6.0% and polymorph Y containing 1.0%-2.0% water are obtained.

[0010] The known polymorphic modifications of rifaximin are crystal hydrates and can transform into each other depending on the water content and temperature - these processes are described in detail in “Crystal forms of rifaximin and their effect on pharmaceutical properties” July 2008 CrystEngComm 10(8). Thus, rifaximin a, when moistened, transforms into the 8 form, which, giving off or accepting moisture, transforms into the £ or (3 form, and p, respectively, depending on the water content and temperature, transforms into the y, s, £ or a modification.

[0011] Patent CN 103509038 describes a crystalline modification of rifaximin C, for the production of which raw rifaximin with moisture up to 5.0% is dissolved in alcohol and rifaximin is crystallized by cooling the solution to a temperature of -10-^--30 оC, the obtained crystalline modification has a characteristic diffraction pattern with the values ​​of the peaks of the diffraction angles 20: 5.8°, 6.4°, 7.8°, 8.9°, 9.3°, 11.4°, 11.8 °, 12.2°, 12.6°, 13.8°, 14.8°, 15.7°, 16.5°, 17.0°, 17.9°, 18.8°, 19.3°, 19.9°, 20.9 °, 21.6°, 23.3°, 25.5°, 28.0°. Rifaximin obtained in this way, like the previous modifications, is a crystalline hydrate, i.e. the solution from which it crystallizes contains water introduced during the dissolution of the original raw crystalline hydrate.

[0012] All the described modifications of rifaximin have a common drawback, namely: they can change from one form to another with changes in humidity and temperature, which leads to changes in the shape, size and density of crystals. Over time, pure polymorphic modifications come into equilibrium with other modifications, forming an equilibrium composition characteristic of a certain temperature and humidity, the most energetically favorable for a given thermodynamic state. The occurrence of the listed processes in dosage forms prepared on the basis of rifaximin crystal hydrates can lead to a change in their properties, in particular solubility, bioavailability, and a change in pharmacokinetics. In addition, a change in the shape and size of crystal hydrates of the active pharmaceutical component in solid drugs can lead to a change in the physicomechanical characteristics of the drug.

[0013] Attempts to obtain stable rifaximin crystal hydrates characterized by a constant moisture content are described in Russian patent 2593750 C2. The stability of the polymorphic form is demonstrated by exposing a rifaximin sample to an environment with a relative humidity of 80% for a long time, up to 5 months, during which the moisture content in the rifaximin sample remains virtually unchanged and constant at 4.5%.

[0014] Patent EA 030457 B1 describes a rifaximin solvate T, which contains rifaximin and diethylene glycol monoethyl ether in a stoichiometric ratio with rifaximin of 1:1. According to the description, this form of rifaximin does not contain water and does not change its structure when exposed to moisture for 10 days. The main disadvantage of this form of rifaximin is the presence of diethylene glycol monoethyl ether, the content of which, at the specified molar ratio, is 10.3%. Ethylene glycol monoethyl ether, in accordance with the classification of pharmaceutical toxicity according to ICH guideline Q3C (R6) of the European Medicines Agency, belongs to the second class of toxicity and its content in drugs should not exceed 0.016%; due to its high toxicity, the use of such a rifaximin solvate for pharmaceutical purposes is very limited.

[0015] As the closest analogue of the present invention, one can consider international application WO 2018 / 197538, which discloses a pharmaceutical composition with an antibiotic effect, containing an anhydrous polymorphic form of rifaximin [3 (water content 9.1 wt %), in a therapeutically effective amount. Also, said composition contains microcrystalline cellulose, sodium carboxymethyl starch, talc, silicon dioxide and magnesium stearate in appropriate amounts, while the composition can be in the form of a tablet or capsule. The disadvantage of this pharmaceutical composition is that the water content in the [3 form significantly exceeds 1%, which does not allow to completely exclude the effect of water on bioavailability, the period of the drug in the systemic bloodstream and intestinal lumen, the development of possible resistance of microorganisms, as well as therapeutic efficacy.

[0016] In the cited patent documents, the stability of rifaximin is demonstrated by its resistance to moisture, however, the tests conducted do not fully characterize the stability of the polymorphic form, since in fact, modern tablet dosage forms are stored in hermetically sealed containers, usually in closed glass jars or blisters, such packaging ensures constant humidity of the environment with which the tablet form comes into contact, and the main variable factor affecting the drug is temperature, the fluctuations of which are associated with both daily changes day - night, and seasonal summer - winter. Temperature fluctuations lead to fluctuations in the thermodynamic equilibrium established between the various crystal hydrate structures of rifaximin, which further lead to fluctuations in the shape and size of the crystals and the accumulation of deformations in the tablets.Changes in the polymorphic composition of the active substance and the parameters of the drug, such as tablet strength and its disintegration, cause changes in the pharmacokinetic properties of the drug, reduce the predictability of its effect on the body and reduce the effectiveness of treatment.

[0017] The effectiveness and safety of rifaximin in the treatment of intestinal infections is due to its extremely low solubility in water, which makes it virtually impermeable through the intestinal walls. When taken orally, only about 1% of rifaximin enters the blood, so its effect on internal organs is minimal, and 99% of rifaximin is distributed throughout the intestine, providing high activity inside the esophagus. The solubility of the substance is determined by its polymorphic form, and the release of the substance from the tablet and the specified distribution along the esophagus depend on the properties of the tablet, so the determining factors for the stable effectiveness and safety of rifaximin are the stability of the polymorphic form of rifaximin and the stability of the physicochemical properties of the tablet.

[0018] Brief summary of the invention

[0019] The aim of the present invention is to create a non-toxic pharmaceutical composition of rifaximin with high physicochemical and pharmacokinetic stability over wide time and temperature ranges and a prolonged effect.

[0020] The said objective is achieved in that the claimed pharmaceutical composition, which has an antibiotic effect, contains a therapeutically effective amount of crystalline rifaximin taken in anhydrous form, and also contains, per one weight part of rifaximin, microcrystalline cellulose from 0.6 to 0.8 parts, sodium carboxymethyl starch from 0.035 to 0.045 parts, talc from 0.02 to 0.03 parts, silicon dioxide from 0.015 to 0.02 parts, and magnesium stearate from 0.005 to 0.01 parts.

[0021] In a preferred particular case, the pharmaceutical composition comprises rifaximin with a water content of less than 1.0%.

[0022] The pharmaceutical composition according to the invention can be made in the form of a tablet or capsule.

[0023] The studies conducted by the authors showed that the rate of transition of some polymorphic forms of rifaximin to others, in equilibrium with them, is determined by the moisture content in the rifaximin crystal hydrate, so the highest rate of crystal transformation is observed at a moisture content of 3.0%-6.0%. Oscillatory changes in temperature from -30°C to +60°C in samples with moisture of 3.0%-6.0% after 14 heating and cooling cycles lead to the appearance of both individual pink crystals and light streaks throughout the entire volume of the powder in the dark red powder, which amounted to 3% to 5% of the powder mass. After 60 cycles of temperature oscillations, the inhomogeneity of the powder already amounted to 25% to 30% of the powder mass.

[0024] A decrease in moisture content to 1.0% - 3.0% leads to a sharp decrease in the rate of transition of polymorphic forms to different equilibrium states caused by temperature fluctuations, and the transformation of crystals slows down. Cyclic temperature changes after 14 cycles do not disrupt the homogeneity of the powder, signs of heterogeneity appear only after 60 cycles of thermal fluctuations.

[0025] It was also unexpectedly discovered that with a further decrease in water content to 1.0% or less, the transitions of rifaximin into other polymorphic modifications cease, and the crystal shape and color of the crystals remain constant. An active pharmaceutical substance with such properties is the most suitable for obtaining drugs with a predictable biochemical effect over a long shelf life. Dosage forms containing rifaximin with moisture less than 1.0% retain high stability of both physicochemical and pharmacological properties, which makes them more convenient both for the production of pharmaceuticals and for their storage and transportation.

[0026] The invention can be implemented by obtaining rifaximin with a water content of less than 1.0% by crystallization from a dried solution of raw rifaximin, thereby eliminating the possibility of forming crystal hydrates and the resulting rifaximin containing only one anhydro polymorphic modification. The most suitable solvents are methyl, ethyl and propyl alcohols. Crystallization is carried out by dissolving raw rifaximin, with a content of the main substance of about 90% and moisture of 5-10%, in one of the listed alcohols or a mixture thereof. For dissolution, 2 to 10 parts of solvent are loaded per one part of rifaximin, the ratio is selected for a specific process depending on the dissolution temperature, viscosity of the resulting solution and the residual concentration of the target substance in the mother liquor. Rifaximin is dissolved by heating the suspension to a temperature of 40°-70°C.The resulting solution is dehydrated using inert sorbents, the most suitable are calcium chloride, sodium sulfate and copper sulfate, or moisture is removed by azeotropic distillation of water. The dehydrated solution is cooled until crystallization begins or it is provoked by adding seed crystals. The mixture is stirred for 2-5 hours until a stable suspension is formed and cooled to a temperature of +5° - 0°C, kept at this temperature for 8-16 hours until the crystallization process is complete, filtered and washed with cooled alcohol, the paste is dried from residual amounts of solvent. The crystalline rifaximin isolated in this way contains 98.0%-99.5% of the main substance and 0.2%-0.6% water. The diffraction pattern of the obtained rifaximin with a Cu emitting tube has characteristic maxima of diffraction peak angles of 26: 5.12°; 5.35°; 7.04°; 8.41°; 8.73°; ​​12.54°; 13.51°; 14.57°; 17.95°; 19.79°; 20.67°; 21.47°; 22.1 G; 23.25°.

[0027] The resulting crystalline rifaximin does not contain crystal hydrate water and therefore does not transform into various equilibrium crystal hydrate modifications, which allows it to remain in an unchanged crystalline form at temperature fluctuations from -30°C to +60°C. The active substance in this form is most suitable for the production of solid dosage forms characterized by high stability of physicochemical and pharmacological properties.

[0028] The composition of rifaximin should also ensure high stability of the tablet throughout the shelf life in a wide range of temperature fluctuations and uniform release of the active pharmaceutical substance in the intestine.

[0029] As has been established and as will be shown below, the above objectives are best achieved in a composition in which, per one part of rifaximin, there is a binder - fine crystalline cellulose, the content of which varies within the range from 0.6 to 0.8 parts by weight, a disintegrant sodium carboxymethyl starch from 0.035 to 0.045 parts by weight and auxiliary substances talc from 0.02 to 0.03 parts, silicon dioxide from 0.015 to 0.02 parts and magnesium stearate from 0.005 to 0.01 part per one part by weight of rifaximin.

[0030] It was also unexpectedly discovered that crystalline rifaximin in the form of anhydroform in the composition of the above-described pharmacomposition shows a lower concentration of rifaximin in plasma in the most important time period corresponding to Tmax = 2-3 hours, which should be attributed to the safety profile of the drug, and also shows a higher concentration of rifaximin in the intestinal lumen (intestinal tissue) in the most important time period corresponding to 2-8 hours, which should be attributed to the effectiveness of the drug, as well as a decrease in the risk of resistance development, showing signs of prolonged action in the finished dosage form.

[0031] All technical and special terms used in the description have the generally accepted meaning in this field of technology.

[0032] Brief description of the drawings

[0033] The present invention is further illustrated by means of Fig. 1 and Fig. 2. Fig. 1 shows diffractograms of a sample of the anhydro form of rifaximin: a - untreated, b - with signed d values ​​for each of the peaks, c - with signed 20 angle values ​​for each of the peaks with the exclusion of the a1+a2 doublet lines. Fig. 2 shows the general scheme of experiments on changing the concentration of rifaximin in the studied blood serum samples over time.

[0034] Detailed description of the invention

[0035] The invention is illustrated by the following illustrative examples, which do not limit the claimed scope of protection. Example 1. Obtaining anhydrous form of rifaximin.

[0036] Propanol (1200 ml) is poured into a 2000 ml flask equipped with a distillation condenser and a stirrer, 365 g of raw rifaximin is added while stirring, with a water content of 4%, the suspension is heated to a boiling point of 88 ° C, during which rifaximin dissolves to form an intensely red solution. Then 200 ml of water-alcohol azeotropic mixture are distilled into a receiving flask. The solution is cooled to a temperature of 15 ° C and seed crystals are added with vigorous stirring, stirring is continued until a stable suspension is formed for 2 hours, after which it is cooled to a temperature of + 3 ° C, and maintained at this temperature for 10 hours. The suspension is filtered, washed with 450 ml of cooled propanol. Dry in a nitrogen stream at a temperature of 50 ° C for 6 hours. The weight of the obtained dry crystalline powder is 299.0 g, the content of the main substance is 99.2%, the water content according to the Fisher method is 0.3%.The parameters of the diffraction pattern lines of the rifaximin sample are presented in Table 1, and the X-ray diffraction pattern of the sample of the obtained anhydrous form of rifaximin is presented in Fig. 1.

[0037] Table 1. Parameters of the diffraction lines of the rifaximin sample

[0038]

[0039] Analysis of the polymorphic modification by the refractoscopy method shows characteristic maxima of the diffraction peak angles 20: 5.12°; 5.35°; 7.04°; 8.41°; 8.73°; ​​12.54°; 13.51°; 14.57°; 17.95°; 19.79°; 20.67°; 21.47°; 22.11°; 23.25°. Example 2. Obtaining the anhydro form of rifaximin.

[0040] A 2000 ml flask is charged with 1400 ml of methanol, 300 g of rifaximin is added, the content of the main substance is 97% and 6% of water, 600 g of anhydrous copper sulfate is added, the suspension is heated to a temperature of 50 ° C and stirred for 12 hours. The dried rifaximin solution is filtered, cooled to a temperature of -10 ° C, the precipitated rifaximin crystals are filtered, washed with 100 ml of cooled methanol. The powder is dried from residual amounts of solvent in a nitrogen stream at a temperature of 50 ° C for 4 hours. The weight of dry crystalline powder is 265.1 g. The content of the main substance is 98.5%, the water content by the Fisher method is 0.25%. The X-ray powder diffraction (XRPD) pattern of the product is characterized by the same 20 values ​​for the diffraction peaks shown in Fig. 1 and corresponds to the X-ray diffraction pattern in Fig. 1 calculated from the data for the single crystal according to Example 1.

[0041] Example 3. Obtaining the anhydrous form of rifaximin.

[0042] In a 2000 ml flask equipped with a reflux condenser, 1000 ml of ethanol are loaded, 160 g of raw rifaximin are added, the suspension is heated to 60 °C, stirred until dissolved, 320 g of anhydrous calcium chloride are added, stirred for 12 hours, the moistened calcium chloride is filtered, the transparent solution is cooled to 5 °C, the precipitated rifaximin is filtered, washed with alcohol, and dried from the solvent. Upon completion of drying the rifaximin, crystalline rifaximin 112+.6 g is obtained, the content of the main substance is 98.5% and moisture is 0.25%. The X-ray powder diffraction (XRPD) pattern of the product is characterized by the same 20 values ​​for the diffraction peaks shown in Fig. 1, and corresponds to the X-ray diffraction pattern in Fig. 1, calculated on the basis of data for a single crystal in accordance with Example 1.

[0043] Example 4. Obtaining a pharmaceutical composition of rifaximin in anhydrous form.

[0044] Rifaximin, obtained as described in example 3 in the amount of 105.5 g, is mixed with 80 g of fine-crystalline cellulose, 1.8 g of silicon dioxide, 4.5 g of sodium carboxymethyl starch, 2.7 g of talc and 0.9 g of magnesium stearate. The mixture is thoroughly mixed until a homogeneous mass is obtained. The mixture is poured onto trays, the height of the mixture layer is about 1 cm, the trays are placed in a vacuum drying cabinet. Dry at a temperature of 65 ° C and a vacuum of 1 mm Hg for 16 hours. At the end of the process, a mass with a moisture content of 0.9% is obtained. Tablets weighing 380 mg, cylindrical in shape and 11 mm in diameter, are pressed from the mass. The resulting tablets are distinguished by high physicochemical stability with temperature fluctuations from -30 ° C to 65 ° C.

[0045] Example 5. Comparison of the properties of rifaximin tablets produced from rifaximin samples with different moisture contents.

[0046] Rifaximin crystallohydrate powder with 5.0% moisture, rifaximin powder with 2.0% moisture obtained by drying rifaximin with 5.0% moisture and rifaximin powder with 0.3% moisture obtained according to Example 1 are used to obtain tablets of the same shape and formulation, for which 100 g of rifaximin are mixed with 80 g of fine-crystalline cellulose, 1.8 g of silicon dioxide, 4.5 g of sodium carboxymethyl starch, 2.7 g of talc and 0.9 g of magnesium stearate. The mixtures of the components are homogenized to obtain uniform tablets of the mass. Cylindrical tablets weighing 380 mg and 11 mm in diameter are pressed from the tablets of the mass. The resulting tablets were sealed in hermetically sealed containers and placed in a thermostat with a cyclically changing temperature from -30°C to 65°C, with the heating cycle lasting 8 hours and the cooling cycle lasting 16 hours. After a certain number of cycles, the tablets were tested for appearance and mechanical strength by compression in the diametrical direction measured in Newtons.The measurement results are summarized in Table 2. Table 2. Dependence of rifaximin tablet characteristics on moisture content with temperature fluctuations.

[0047] The presented results show that the pharmaceutical composition of rifaximin in anhydroform with moisture less than 1% ensures high stability of the dosage form with temperature fluctuations in a wide range. Example 6. Change in the concentration of rifaximin in the studied blood serum samples over time. Outbred, sexually mature male Wistar rats were used, 30 individuals per group. The number of groups was 3: the first group received the test drug characterized in the formula of the present invention; the second group received the reference drug (Alfa-Normix, Italy, LP-008488), the third group was the control group (water orally). The animals were obtained from the laboratory animal nursery "Rappolovo" and during the study were kept in the vivarium premises under controlled conditions: ambient air temperature 22 ± 2°C, relative humidity 60 ± 5%, with a natural change in the daily cycle and with free access to complete feed and water.Animal care and handling complied with the provisions of Directive EU 2010 / 63 "On the protection of animals used for scientific purposes". Animals were orally administered rifaximin and the reference agent once at a dose of 418 mg / kg (calculated taking into account the interspecies dose conversion factor and the daily dose of rifaximin of 2400 mg). Blood was collected from the animals at time intervals (0.5 h, 1 h, 2 h, 4 h, 8 h, 16 h, 24 h, 48 h), followed by obtaining serum and determining the concentration of rifaximin in the blood according to the method of the current pharmacopoeial article 1. Blood was collected from the sublingual vein using castaviejo scissors into citrate-filled Eppendorf tubes. The volume of blood collected at each stage was 0.35 ml. The whole blood was then centrifuged at 3500 to obtain serum, which was frozen until analysis. The serum was transferred to centrifuge tubes, the proteins were precipitated, and the serum was centrifuged again.The final volume of the analyte was 0.7 ml. Small intestine was collected after cervical dislocation of animals under chloral hydrate anesthesia (350 mg / kg, intraperitoneally). A fragment of the small intestine (L = 10 mm) was washed from the contents with a PBS buffer solution (pH = 7.4), after which it was homogenized in the same solution in a tissue mass:buffer ratio of 1:10. The resulting homogenate was centrifuged at 10,000 g for 20 min. The supernatant was transferred to Eppendorf tubes and used for analysis.

[0048] The content of rifaximin in the test samples was assessed by HPLC. Chromatography conditions: Mobile phase: Acetonitrile - methanol 1:1; column: Agilent ZORBAX; column temperature 40 °C; flow rate 1.4 ml / min; spectrophotometric detector, 276 nm; sample volume 20 μl; chromatograph: "Dionex" UltiMate 3000. The results obtained were processed using variation statistics methods.

[0049] The statistical analysis was performed using the StatPlus 7.0 software package. The normality of data distribution was assessed using the Shapiro-Wilk test. Homogeneity of variances was determined using the Levene test. Statistically significant variables were assessed using the one-way ANOVA with the Newman-Keuls posttest (for normal data distribution) or the Kruskal-Wallis pot test (for data distribution different from normal) at a critical significance level of p<0.05. Paired comparisons were performed using the Mann-Whitney test at p<0.05.

[0050] The change in the concentration of rifaximin mg / ml in the studied blood serum samples over time is presented in Table 3.

[0051] Table 3. Changes in the concentration of rifaximin mg / ml in the studied blood serum samples over time (M±SD)

[0052] Note: * - reliable relative to the 0.5 hour indicator

[0053] (ANOVA, with Newman-Keuls post-processing); ■# - reliable relative to the comparison sample (Mann-Whitney criteria, paired comparison) In the blood plasma samples of rats that received the comparison sample, rifaximin was detected in the time interval of 16 hours at a concentration of ~ 4% of the administered dose. Moreover, starting from the 4th hour, a statistically significant decrease in the content of rifaximin in the blood plasma of rats was observed in relation to the indicator obtained after 0.5 hours after the administration of rifaximin. In animals that were administered the test sample, rifaximin was detected in the blood plasma in the time interval of 24 hours at a concentration of ~ 3% of the administered dose. It should be noted that 0.5 hours, 1 hour and 2 hours after the administration of rifaximin, its content in the blood plasma samples of animals that received the test sample was significantly lower than that in rats that were administered the comparison sample.Subsequently, in the interval of 9-16 hours, the concentration of rifaximin in the blood of rats that received the test sample was higher than that in animals that were administered the reference. In the time interval of 0.5-2 hours, a smaller amount of rifaximin was detected in the blood plasma samples of rats that received the test sample than in the plasma of rats that were administered the reference, which may indicate lower systemic absorption of the drug and its presence in the intestinal lumen in higher concentrations.

[0054] Example 7. Changes in the concentration of rifaximin in the studied blood serum samples over time

[0055] Outbred, sexually mature male Wistar rats were used, 30 individuals per group. There were 3 groups: the first group received the test drug described in the formula of the present invention; the second group received the reference drug (Alfa-Normix, Italy, LP-008488), the third group was the control (water orally). The animals were obtained from the Rappolovo laboratory animal nursery and were kept in the vivarium premises under controlled conditions for the duration of the study: ambient air temperature 22±20C, relative humidity 60±5%, with a natural change of the daily cycle and free access to complete feed and water. The maintenance and manipulations with the animals complied with the provisions of Directive EU 2010 / 63 "On the protection of animals used for scientific purposes". The animals were orally administered rifaximin and the reference at a dose of 418 mg / kg (calculated taking into account the interspecies dose conversion factor and a daily dose of rifaximin of 2400 mg).Small intestine was collected after cervical dislocation of animals under chloral hydrate anesthesia (350 mg / kg, intraperitoneally). A fragment of small intestine (L= 10 mm) was washed from contents with PBS buffer solution (pH=7.4), after which it was homogenized in the same solution in a tissue mass:buffer ratio of 1:10. The resulting homogenate was centrifuged at 10,000 g for 20 min. The supernatant was transferred to Eppendorf tubes and used for analysis.

[0056] The content of rifaximin in the test samples was assessed by HPLC. Chromatography conditions: Mobile phase: Acetonitrile - methanol 1:1; column: Agilent ZORBAX; column temperature 40 °C; flow rate 1.4 ml / min; spectrophotometric detector, 276 nm; sample volume 20 μl; chromatograph: "Dionex" UltiMate 3000. The results obtained were processed using variation statistics methods.

[0057] The statistical analysis was performed using the StatPlus 7.0 software package. The normality of data distribution was assessed using the Shapiro-Wilk test. Homogeneity of variances was determined using the Levene test. Statistically significant variables were assessed using the one-way ANOVA with the Newman-Keuls posttest (for normal data distribution) or the Kruskal-Wallis pot test (for data distribution different from normal) at a critical significance level of p < 0.05. Paired comparisons were performed using the Mann-Whitney test at p < 0.05.

[0058] Analyzing the change in the concentration of rifaximin in the intestinal wall, it was shown that in rats administered the reference substance, the content of rifaximin decreased over time (p < 0.05). At the same time, stable concentrations were recorded in the range of 0.5-2 hours. At the same time, in animals that received the test sample, rifaximin was detected in the intestinal wall 24 hours after administration. In the range of 0.5-2 hours, the concentration of rifaximin in the intestinal wall of rats that received the test sample was higher than that in animals that were administered the reference substance, but statistical significance was not recorded (see Table 4).

[0059] Table 4. Changes in the concentration of rifaximin mg / ml in the studied intestinal supernatant samples over time (M±SD).

[0060] The increased content of rifaximin in rats that received the test sample in the interval of 4-24 hours may indicate a slower clearance of rifaximin and its prolonged presence in the intestinal lumen. Also, a slower clearance of rifaximin of the test sample may be indicated by its longer presence in the intestinal wall of rats. An increase in the concentrations of rifaximin in the intestinal lumen of the test drug, against the background of the compared drug, may have important clinical significance, both in terms of increasing its antimicrobial effectiveness and reducing the likelihood of the formation of resistant pathogenic strains.

Claims

Invention formula 1. A pharmaceutical composition characterized by prolonged antibiotic action in the intestinal lumen, containing a therapeutically effective amount of crystalline rifaximin with a water content of less than 1.0% with characteristic diffraction angle maxima 20: 5.12°; 5.35°; 7.04°; 8.41°; 8.73°; 12.54°; 13.51°; 14.57°; 17.95°; 19.79°; 20.67°; 21.47°; 22.11°; 23.25° based on one part by weight of rifaximin: microcrystalline cellulose - from 0.6 to 0.8 parts, sodium carboxymethyl starch from 0.035 to 0.045 parts, talc from 0.02 to 0.03 parts, silicon dioxide from 0.015 to 0.02 parts and magnesium stearate from 0.005 to 0.01 parts.

2. A pharmaceutical composition according to item 1, characterized in that it is made in the form of a tablet or capsule.

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