EA023814B1 - METHOD OF OBTAINING POLYMERIC COMPOSITIONS BASED ON PROANTOCYANIDINE FOR PHARMACEUTICAL FORMS - Google Patents

Abstract

Methods for the preparation of pharmaceutical compositions containing proanthocyanidin polymer used for the treatment and prevention of various conditions are proposed. In particular, the invention relates to a process for the preparation of pharmaceutical forms of a proanthocyanidin-based polymer composition, which has been isolated from Croton spp. or Calophyllum spp. In particular, the invention relates to a method for producing proanthocyanidin-based polymer compositions that achieve concentrations and degrees of purity suitable for addition to pharmaceutically effective forms.

Description

Secretory diarrhea, also called water diarrhea, is the leading cause of morbidity and mortality in developing countries, especially infants and young children. Secretory diarrhea also affects a significant portion of visitors from developed to developing countries, but can affect anyone when visiting a foreign country (the so-called diarrhea of travelers). Secretory diarrhea is characterized by loss of fluid and electrolytes through the intestinal tract, leading to severe and sometimes life-threatening dehydration. Secretory diarrhea is caused by various bacterial, viral and protozoal pathogens, and also has a different, non-infectious etiology, such as cancerous tumors and tumors of the gastrointestinal tract.

The two main bacterial sources of secretory diarrhea are UShtyu syo1egae (U.Syoegae) and Exxepsoy soi (E.soy). Enterotoxigenic E. soya species are a source of secretory diarrhea in developing countries and are the main cause of travelers' diarrhea. Other E. coli strains that cause diarrhea include enterohemorrhagic, enteroinvasive and enteropathogenic and other strains. Other bacterial agents that cause diarrhea include other UVtu crp., Satru1baclcr. , VasYupobek krr., 81aryu1osossik krr. and VasShik krr., as well as other intestinal bacteria. Secretory diarrhea can also be caused by protozoal pathogens, such as SgurYukropbsht krr., For example Sguryukropbsht staguit. See mainly No11aib, 1990, Sy. Muto Kyu. 3: 345; Natk, 1988, App. Sue. Bb. 8 18: 102; Sgaseu, 1986, Sy. ίη Sak1toi1., 15:21; Ootk and Oedtuk, 1986, Beetattu Kek. Sott. 10: 355; B1ask, 1982, Meb. Sue. Yog. At. 66: 611.

U. syo1egae, enterotoxigenic E. coli strains and a number of other intestinal bacteria cause secretory diarrhea through similar mechanisms. These pathogens produce a toxin that binds to a specific receptor on the apical membrane of the intestinal epithelium. Receptor binding initiates transduction of the signal mediated by adenylate cyclase or guanylate cyclase, leading to an increase in the amount of cAMP or cGMP. The indicated regulatory cascade, apparently acting by phosphorylation of specific proteins of the apical membrane, stimulates the outflow of chloride into the intestinal lumen from intestinal crypt epithelial cells and inhibits the normal resorption of sodium and chlorine ions through intestinal villous epithelial cells. An increased concentration of chloride and sodium ion osmotically draws water into the intestinal lumen, which leads to dehydration and loss of electrolytes. Agents that reduce the secretion of chloride ion, therefore, will interfere with the movement of fluid in the intestine and, as a result, the excretion of fluid. Thus, such agents are especially useful in the treatment and prevention of dangerous dehydration and loss of electrolytes caused by secretory diarrhea.

Secretory diarrhea is also a serious problem for non-human animals, especially farm animals such as cattle, pigs, sheep (sheep and rams), birds (such as chickens) and horses, and other domesticated animals such as dogs and cats. Diarrhea is especially common in young and recently weaned farm animals. Diarrhea in farm animals, especially beef and dairy cattle, such as cattle, sheep, and pigs, is often caused by bacterial pathogens such as enterotoxigenic, enterohemorrhagic, and others E. coli, Sariconida regensis, Cactus sylvestris, Basqueoccus. Sashru1oBas1et krr. and the wacky girl. In addition, protozoal pathogens, especially SturFrkrotSht ratit, and viral agents, especially rotaviruses and coronaviruses, are the main sources of diarrhea in farm animals. Other viral agents that cause diarrhea in farm animals include togavirus, parvovirus, calcivirus, adenoviruses, bredaviruses, and astroviruses. See mainly No11aib, 1990, Sy. M1stoYu1o§u Keu. 3: 345; see also Sy / \\ Sheg and Blit, 1996, A1UK 57: 560; Shtotesk, 1995, Uletshatu Siag1eg1u 17 (adj. 1): W12; Uetti, 1994, Aik1ta1. Ueetsatu 1. 71:33; Guardians and others, 1993, Aik1ta1. Step1: 70: 259; Moshtsoy, 1991, Eig. 1. YerShetyuy 7: 588; Ootk and Oedguke, 1986, Beetle Step Kek. Sosh. 10: 355.

Proanthocyanidin and proanthocyanidin polymers are phenolic substances found in a wide variety of plants, especially those with a woody nature of development (for example, Stoelo crp. And Cauloptera crp.). The general chemical structure of polymeric proanthocyanidin consists of linear chains that contain 5,7,3 ', 4'-tetrahydroxy or 5,7,3', 5'-pentahydroxyflavan-3-sex units, interconnected via C (4) - ( 6) and / or C (4) -C (8).

Biosynthetic studies have shown that proanthocyanidin polymers are composed of monomer units. See P1e1set et al., 1977, 1.S. Sh. Regksh, 1: 1628. The monomer unit (mainly called leukoanthocyanidin) of the polymer chain can be based on one of two isomers with stoichiometry in the C-ring, at position 2 and / or 4, referred to as cis (the so-called epicatechins) or trans (the so-called catechins). Thus, polymer chains are based on various structural units, which cause a wide variety of polymer proanthocyanidins and a large number of possible isomers (Nepyi, yau et al., 1982, 1. S.S. Retkt, 1: 1217). ¹³ C NMR spectroscopy was used to determine the structures of polymeric proanthocyanidins, and recent work has shed light on the chemical composition of di-, tri- and tetrameric proanthocyanidins. In most plants, 1,023,814 plants are dominated by larger polymers with flavan-3-tin units with an average molecular weight of more than 2,000 Da containing 6 or more units (No. utup et al., 1987, Mad. Kek. Sit., 25: 118).

SUMMARY OF THE INVENTION

The invention provides methods for producing proanthocyanidin-based polymer compositions for use in pharmaceutically effective dosage forms. In particular, the invention provides methods for the preparation of pharmaceutical forms of polymer compositions based on proanthocyanidin that have been isolated from Crgopcr. or Ca1 / o11it crp., these forms can be effective, for example, for the treatment of secretory diarrhea, in particular to reduce fluid loss and the resulting dehydration, which are associated with secretory diarrhea. Methods are provided for preparing a proanthocyanidin polymer that is suitable for pharmaceutical forms having a high level of purity and a therapeutically effective concentration.

The invention relates to methods for producing a polymer composition based on proanthocyanidin, which has a degree of purity and concentration suitable for use in therapeutically effective pharmaceutical compositions. A polymer composition based on proanthocyanidin can be obtained, for example, from the milk juice CrOiop crr., Especially Croiop 1ecyiep. The proanthocyanidin polymer in the composition obtained according to the methods proposed in the present invention is crofelemer (ϋδΑΝ). Also described in this application are new pharmaceutical compositions prepared according to a method for producing a polymer composition based on proanthocyanidin. The methods described herein provide proanthocyanidin-based polymer compositions containing more than 90 wt.% Proanthocyanidin polymer (the chromatographic purity of which can be determined, for example, by analysis of the components to be determined compared to a standard sample using chromatography), more than 95 wt.% proanthocyanidin polymer and or, for example, from about 95 to about 100 wt.% proanthocyanidin polymer. The present invention also provides polymer compositions based on proanthocyanidin containing less than about 0.2% taspine (the chromatographic purity of which is determined by analysis of the determined components compared to a standard sample using chromatography). In addition, the invention also provides polymer compositions based on proanthocyanidin having increased uniformity. For example, the proanthocyanidin-based polymer compositions of the present invention have a polydispersity value ranging between about 1.2 and about 1.8, or, for example, between about 0.9 and about 1.2, or from about 0 5 to about 1.5, or between about 0.8 and about 1.3.

In one embodiment of the invention, a method for producing a polymer composition based on proanthocyanidin is provided, which comprises (ί) keeping the milk juice CrClop crr. at a temperature below room temperature for at least 48 hours to allow sedimentation of suspended particles; (ίί) adjusting the pH of the milky juice to a value in the range of from about 6.5 to about 8.5; (ίίί) filtering the milky juice to obtain a filtrate; (ίν) adjusting the pH of the filtrate to a value in the range of from about 3.5 to about 5.5; (ν) performing solid phase extraction of the filtrate to obtain an eluate containing a proanthocyanidin polymer; (νί) performing solid phase extraction with a solid phase different from the solid phase in step (ν), to obtain an eluate containing a proanthocyanidin polymer and (νίί) processing the eluate to obtain a polymer composition based on proanthocyanidin. The method may include an additional, optional step, which is an additional processing step similar to step (νίί) but located between steps (ν) and (νί).

Alternatively, a method for producing a proanthocyanidin-based polymer composition may include providing or preparing a partially purified proanthocyanidin-based polymer composition; performing extraction (eg, solid phase extraction) of a partially purified proanthocyanidin-based polymer composition to obtain an eluate containing a proanthocyanidin polymer; and treating the eluate to obtain a purified proanthocyanidin-based composition. In one embodiment of the invention, the extraction is carried out using a solid phase extraction resin, such as ion exchange resins, adsorption resins, separation resins, size exclusion chromatography resins, mixed resin and mixtures of more than one type of resin.

In another embodiment, a method for producing a proanthocyanidin polymer comprises:

a) providing a solution of milky juice obtained from a C'goUp 1Syep plant (for example, crude plant milky juice, partially purified plant milky juice, concentrated crude plant milky juice or concentrated partially purified plant milky juice) containing a precipitate;

b) adding methyl ethyl ketone to the plant’s milky juice solution and sediment;

c) separating the organic layer and concentrating the aqueous layer to obtain a solid;

as an alternative or as an alternative, separating the aqueous layer and concentrating the organic layer to obtain a solid;

ά) dissolving a solid in an aqueous solvent;

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e) chromatography of the solution using an eluting solvent; and e) isolation of proanthocyanidin containing Crofelemer.

In the present invention, a precipitate means a suspension obtained during storage. The crude milky sap of the plant can be obtained from the bark of Sto1oi 1eeNep. The specified milky juice is collected and stored in barrels at a temperature of 0-2 ° C. Suspended particles deposited during storage are called sediment. The specified precipitate is mainly removed.

The present invention provides methods for producing a polymer composition based on proanthocyanidin, in some embodiments of the invention, this method may include additional processing by coprecipitation of impurities from a solution containing a polar solvent, further including filtering the material (for example, use a methanol solution with phyllosilicate (for example, one or more with material including bentonite, prenite, clay minerals, palygorskite, muscovite, clintonite, etc. and diatomite, such as commercially available under the names Ce1a1ot® or CeSe®, cellulose mesh filter media, perlite, charcoal, glass filter element, sand, filter paper, Oo \ ye, salt or similar materials) as a filter material and acetate as solvent (e.g. methyl acetate, ethyl acetate)) during extraction and / or washing. For example, filter materials, such as phyllosilicate, can help remove impurities, such as taspine, protein, and other substances. Acetate can help dissolve and subsequently remove leachable materials, such as low molecular weight related substances or low molecular weight phenolic compounds. Additional processing may occur before, after or instead of solid phase extraction. Alternatively, a method for producing a proanthocyanidin-based polymer composition may include purification methods known in the art, and also includes dissolving the partially purified milky juice in alcohols (e.g., methanol, ethanol, propanol, butanol and other alcohols) and extracting the dissolved compound using alcohol, followed by treatment with charcoal and filtration through bentonite. The filtrate in some embodiments of the invention can be further treated with an acetate solvent, followed by filtration through a dense filter (for example, paper, for example, HUNaCap) to obtain the desired compound that does not contain taspine and other impurities.

In addition, the methods proposed in the present invention include methods that are suitable for industrial production. For example, the methods proposed in the present invention can be applied to minimize the need for the disposal of toxic materials, solvents, and / or can be applied to reduce the volume of liquid at a certain stage of the method on an industrial scale.

In another embodiment, a pharmaceutical form comprising a proanthocyanidin-based polymer composition is provided. Preferably, the pharmaceutical form is intended for oral use.

According to one aspect, the present invention provides methods for producing a pharmaceutically acceptable polymer composition based on proanthocyanidin, including (ί) providing for the sedimentation of particles suspended in the milk juice of CGyOp 8 pp; (ίί) adjusting the pH of the milky juice to a value in the range of from about 6.5 to about 8.5; (ίίί) filtering the milky juice to obtain a filtrate; (ΐν) adjusting the pH of the filtrate to a value in the range of from about 3.5 to about 5.5; (ν) performing a first extraction of the filtrate to obtain an eluate containing a proanthocyanidin polymer; and (νί) collecting an eluate containing a proanthocyanidin polymer composition.

In one embodiment of the invention, the first extraction comprises solid phase extraction.

In one embodiment of the invention, the method further comprises performing a second extraction, wherein the second extraction comprises solid phase extraction.

In one embodiment of the invention, the second solid phase is different from the first solid phase.

In one embodiment, the proanthocyanidin polymer composition comprises more than 90% of the proanthocyanidin polymer.

In one embodiment, the proanthocyanidin-based polymer composition comprises more than 95% proanthocyanidin polymer.

In one embodiment of the invention, the proanthocyanidin-based polymer composition mainly comprises a proanthocyanidin polymer and water.

In one embodiment, the suspended particles are allowed to settle for at least 48 hours.

In one embodiment of the invention, the suspended particles are allowed to settle over a period of time ranging from about 48 hours to about 48 months.

In one embodiment of the invention, the suspended particles are allowed to settle at a temperature of less than about 20 ° C; at a temperature between about 20 and about 0 ° C, between about 30 and about 20 ° C, between about 20 and about -20 ° C, or between about 0 and 15 ° C.

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In one embodiment of the invention, the suspended particles are allowed to settle at a temperature between about 0 ° C. and 15 ° C. for at least 2 hours.

In one embodiment, the pH is adjusted to pH 8 in step (s).

In one embodiment of the invention, step (ίίί) is carried out using an auxiliary filter material.

In one embodiment, step (этап) further includes one or more water washes of the filter material.

In one embodiment, step (ίίί) further includes centrifuging the milk juice instead of filtering it.

In one embodiment of the invention, the filter material is one or more materials including diatomaceous earth, charcoal, bentonite, cellulose, glass, sand or filter paper.

In one embodiment of the invention, the filtrate obtained in step (ίίί) and water after any optional washes are combined and treated by ultrafiltration.

In one embodiment of the invention, ultrafiltration is carried out using a semipermeable membrane.

In one embodiment of the invention, the semi-permeable membrane allows the passage of dissolved substances, up to having a molecular weight selected from the group consisting of 500 Da, 1, 5, 10, 20 and 30 kDa.

In one embodiment of the invention, in step (ίν), the pH is 4.

In one embodiment of the invention, in step (ν), solid phase extraction is carried out using a solid phase extraction resin.

In one embodiment of the invention, the solid phase extraction resin is selected from the group consisting of ion exchange resins, affinity resins, adsorption resins, separating resins, and mixtures thereof.

In one embodiment of the invention, the solid phase extraction resin comprises a carboxymethyl modified agarose resin.

In one embodiment of the invention, step (ν) is carried out in batch mode.

In one embodiment of the invention, the method further comprises treating the eluate obtained from the first solid phase extraction to obtain a composition subjected to the second solid phase extraction.

In one embodiment of the invention, the eluate obtained by the first extraction is produced by washing out from the solid phase using a complex of solvents selected from the group consisting of water, acetone, methanol, ethanol, glycol and mixtures thereof.

In one embodiment of the invention, the second solid phase extraction is carried out using a resin for solid phase extraction.

In one embodiment, the solid phase extraction resin is one or more resins, including size exclusion chromatography resins, ion exchange resins, affinity resins, adsorption resins, separating resins, or mixtures thereof.

In one embodiment of the invention, the solid phase extraction resin comprises a modified polysaccharide.

In one embodiment of the invention, the modified polysaccharide comprises hydroxypropylated cross-linked dextran.

In one embodiment of the invention, the eluate obtained by the first extraction is obtained by eluting the solid phase with a solvent selected from the group consisting of water, acetone, methanol, ethanol, glycol and mixtures thereof.

In one embodiment, the eluate obtained from the second extraction is mixed with a solvent selected from the group consisting of methanol, ethanol, propanol, butanol, pentanol, hexanol, ethylene glycol, propylene glycol, ethyl acetate, dichloromethane, trichloromethane, carbon tetrachloride, and dichloroethane and dichloroethane and mixtures.

In one embodiment of the invention, the treatment in step (νί) is selected from the group consisting of ultrafiltration, freeze-drying, evaporation using heat, evaporation without applying heat, evaporation under vacuum, evaporation without applying vacuum, spray drying, and combinations thereof.

In one embodiment of the invention, the level of residual taspine in the proanthocyanidin-based polymer composition is less than 0.2% of the composition.

In one embodiment of the invention, the level of residual taspine in the polymer composition based on proanthocyanidin is less than 0.1% of the amount of composition, the purity of which is determined by chromatography.

In one embodiment of the invention, the level of taspine is determined by chromatography.

In one embodiment of the invention, the polydispersity value of proanthocyanidin is in the range between about 1.2 and about 1.8.

In one aspect, the present invention provides proanthocyanidin-based polymer compositions comprising a proanthocyanidin polymer prepared according to the method described in the description and claims of the present invention.

In one embodiment, the proanthocyanidin polymer composition comprises more than 90% of the proanthocyanidin polymer.

In one embodiment, the proanthocyanidin polymer composition comprises more than about 95% proanthocyanidin polymer.

In one embodiment of the invention, the proanthocyanidin-based polymer composition comprises a proanthocyanidin polymer and water.

In one aspect, the present invention provides pharmaceutical forms comprising a proanthocyanidin polymer composition, the proanthocyanidin polymer composition comprising more than 90% of the proanthocyanidin polymer, and a pharmaceutically acceptable carrier.

In one embodiment of the invention, the level of residual taspine in the proanthocyanidin-based polymer composition is less than 0.2% of the composition.

In one embodiment of the invention, the level of residual taspine in the proanthocyanidin-based polymer composition is less than 0.1% of the composition.

According to one aspect, the present invention provides methods for producing a proanthocyanidin-based polymer composition, comprising (ί) preparing a partially purified proanthocyanidin-based polymer composition, (ΐΐ) performing solid phase extraction of the partially-purified polymer composition to obtain an eluate containing a proanthocyanidin polymer and (ίίί) processing the eluate to obtain a proanthocyanidin composition; moreover, step (ΐΐ) is carried out using a solid-phase extraction resin as a solid phase.

In one embodiment of the invention, the partially purified proanthocyanidin-based polymer composition is partially purified by a method involving the production of milky juice from Cgoca crr. and performing solid phase extraction using a solid phase extraction resin other than hydroxypropylated cross-linked dextran.

In one embodiment of the invention, the solid phase extraction resin comprises a carboxymethyl modified agarose resin.

In one embodiment of the invention, the partially purified proanthocyanidin-based polymer composition is partially purified by a method involving the production of milky juice from Cgoca crr. and performing solid phase extraction using a solid phase extraction resin other than a carboxymethyl modified agarose resin.

In one embodiment of the invention, the solid phase extraction resin comprises hydroxypropylated crosslinked dextran.

In one embodiment, the partially purified proanthocyanidin-based polymer composition comprises from about 35 to about 90% of the proanthocyanidin polymer.

In one embodiment of the invention, a partially purified proanthocyanidin-based polymer composition comprises 8B-300.

In one embodiment of the invention, the solid phase extraction resin used in step (ΐΐ) is a modified polysaccharide.

In one embodiment of the invention, the modified polysaccharide comprises hydroxypropylated cross-linked dextran.

In one embodiment of the invention, the solid-phase extraction resin used in step (ίί) comprises a carboxymethyl-modified agarose resin.

In one embodiment of the invention, the purified proanthocyanidin-based polymer composition comprises more than about 90% proanthocyanidin polymer.

In one embodiment, the purified proanthocyanidin-based polymer composition comprises more than about 95% proanthocyanidin polymer.

In one embodiment, the purified proanthocyanidin polymer composition comprises a proanthocyanidin polymer and water.

In one embodiment, the eluate is mixed with a solvent selected from the group consisting of methanol, ethanol, propanol, butanol, pentanol, hexanol, ethylene glycol, propylene glycol, ethyl acetate, dichloromethane, trichloromethane, carbon tetrachloride, dichloroethane and mixtures thereof.

In one embodiment of the invention, the treatment is selected from the group consisting of ultrafiltration, freeze-drying, evaporation using heat, evaporation without using heat, evaporation under vacuum, evaporation without using vacuum, spray drying, and combinations thereof.

In one embodiment of the invention, the level of residual taspine in the proanthocyanidin-based polymer composition is less than 0.2% of the composition.

In one embodiment of the invention, the level of residual taspine in the purified proanthocyanidin-based polymer composition is less than 0.1% of the composition.

In one embodiment of the invention, the polydispersity of the proanthocyanidin polymer in the proanthocyanidin-based polymer composition is in the range between about 1.2 and about 1.8.

According to one aspect, the present invention provides proanthocyanidin-based polymer compositions comprising a proanthocyanidin polymer obtained according to a method that comprises (ί) preparing a partially purified proanthocyanidin-based polymer composition, (ίί) performing solid phase extraction of a partially purified polymer composition to obtain an eluate, containing a proanthocyanidin polymer and (ίίί) treating the eluate to obtain a proanthocyanidin composition; moreover, step (ίί) is carried out using a solid-phase extraction resin as a solid phase.

In one embodiment of the invention, the proanthocyanidin-based polymer composition has a purity and concentration sufficient to be included in a therapeutically effective pharmaceutical composition.

In one embodiment, the purified proanthocyanidin polymer composition comprises more than 90% of the proanthocyanidin polymer.

In one embodiment, the purified proanthocyanidin-based polymer composition comprises more than 95% proanthocyanidin polymer.

In one embodiment of the invention, the proanthocyanidin polymer composition mainly consists of a proanthocyanidin polymer and water.

According to one aspect, the present invention provides pharmaceutical forms comprising a purified proanthocyanidin polymer composition obtained according to a method which comprises (ί) preparing a partially purified proanthocyanidin polymer composition, (i) performing solid phase extraction of the partially purified polymer composition to obtain an eluate containing a proanthocyanidin polymer and (ίίί) processing the eluate to obtain a proanthocyanidin composition, wherein step (ίί) is carried out with changing the resin for solid phase extraction as a solid phase; and a pharmaceutically acceptable carrier.

In one embodiment of the invention, the level of residual taspine in the purified proanthocyanidin-based polymer composition is less than 0.2% of the composition.

In one embodiment of the invention, the level of residual taspine in the purified proanthocyanidin-based polymer composition is less than 0.1% of the composition.

According to one aspect, the present invention provides methods for producing a proanthocyanidin-based polymer composition, comprising (ί) keeping the Milky Juice 8gp milk juice. at a temperature below room temperature for at least 48 hours to ensure sedimentation of suspended particles and either step (b) (a), or step (s) (b), or step (s) (a) and step (b) (b ) as follows: (i) (a) dissolving the milky juice in water or methanol, adding bentonite to the solution with stirring and optionally adjusting the pH to a value in the range from 5.0 to 7.0 and filtering to obtain a filtrate and / or ( g) (b) adding ethyl acetate and optionally adding water and / or adjusting the pH to a value in the range of 6.5 to 8.5, mixing and removing the ethyl acetate layer atm to obtain an aqueous or methanolic phase or the solid precipitate, and thus, obtaining polymer compositions based proanthocyanidin.

In one embodiment of the invention, the method further comprises purifying the aqueous or methanol phase by phase extraction to obtain a product.

In one embodiment of the invention, the purification comprises solid phase extraction.

In one embodiment of the invention, step (ίί) (a) occurs in methanol.

In one embodiment of the invention, step (ίί) (a) is repeated one to twenty times.

In one embodiment of the invention, step (b) (b) is repeated one to twenty times.

In one embodiment of the invention, in step (ίί) (a), approximately 10% or more of the proteins in the sample are removed prior to step (ίί) (a).

In one embodiment of the invention, the level of taspine in milk juice of more than 250 ppm is reduced to less than about 250 ppm.

According to one aspect, the present invention provides methods for producing a pharmaceutically acceptable polymer composition based on proanthocyanidin, including (ί) ensuring the sedimentation of particles suspended in the milk juice CGUp vr .; (i) adjusting the pH of the milky juice to a value in the range of from about 6.5 to about 8.5; (ίίί) filtering the milky juice to obtain a filtrate; (ίν) adjusting the pH of the filtrate to a value in the range of from about 3.5 to about 5.5; (ν) performing a first solid phase extraction of the filtrate to obtain an eluate containing a proanthocyanidin polymer; and (νί) collecting an eluate containing a proanthocyanidin polymer composition.

According to one aspect, the present invention provides methods for producing a polymer composition based on proanthocyanidin, comprising a) providing a solution of the milky juice obtained from the CgOp 1ceHcg1 plant. containing sediment; B) adding an organic solvent to the plant’s milky juice solution and sediment; c) separating the organic layer and concentrating the aqueous layer to obtain a solid.

In one embodiment of the invention, the method further comprises separating the aqueous layer and concentrating the organic layer to obtain a solid; ά) dissolving a solid in an aqueous solvent; e) chromatography of the solution using an eluting solvent; and ί) isolation of proanthocyanidin containing Crofelemer and, thus, obtaining a polymer composition based on proanthocyanidin.

According to one aspect, the present invention provides methods for producing a proanthocyanidin-based polymer composition, the method comprising: a) providing a solution of milky juice obtained from a plant G1O1E1BE1 containing a precipitate; B) adding an organic solvent to the plant’s milky juice solution and sediment; c) separating the organic layer and concentrating the aqueous layer to obtain a solid or, as an alternative, separating the aqueous layer and concentrating the organic layer to obtain a solid; ά) dissolving a solid in an aqueous solvent; e) chromatography of the solution using an eluting solvent; and ί) isolation of proanthocyanidin containing Crofelemer and, thus, obtaining a polymer composition based on proanthocyanidin.

In one embodiment of the invention, the organic solvent used in step (b) is selected from one or more solvents including alcohols, ketones, esters, ethers, or mixtures thereof.

In one embodiment, the ketone comprises methyl ethyl ketone.

In one embodiment, the organic solvent used in step (b) is selected from a mixture of methanol and ethyl acetate.

In one embodiment of the invention, the column used for chromatography in step (e) comprises a separate column or unit of two columns selected from CM-8percope Cancer! Iote Sulp and Zerbabh HN-20.

Other aspects and embodiments of the invention are provided below.

Brief Description of the Figures

In FIG. 1 shows an amino acid analysis of a sample of a partially purified polymer composition before treatment with bentonite in an aqueous solution;

in FIG. 2 - amino acid analysis of a sample of a partially purified polymer composition after treatment with bentonite in an aqueous solution;

in FIG. 3 is an enlarged chromatogram of the product after treatment with bentonite, coprecipitation with ethyl acetate and first filtration;

in FIG. 4 — amino acid analysis of a sample of a partially purified polymer composition before treatment with bentonite in a methanol solution;

in FIG. 5 - amino acid analysis of a sample of a partially purified polymer composition after treatment with bentonite in a methanol solution;

in FIG. 6 is a flowchart of a method proposed in the present invention;

FIG. 7 is a reverse phase HPLC chromatogram of a product obtained by the method of the present invention using LH-20 resin.

DETAILED DESCRIPTION OF THE INVENTION

The invention provides methods for producing a polymer composition based on proanthocyanidin and a composition obtained according to the specified method. The starting material is the milky juice of the plant, for example, the milky juice of Csoyup crr. or Ca'lbu11it crp. In one embodiment of the invention, the proanthocyanidin-based polymer composition is prepared from the milk juice CGyOp1eErrg. In another embodiment, the proanthocyanidin-based polymer composition is obtained from the milk-based juice of Canthomonitum toribut. In one embodiment of the invention, the proanthocyanidin-based polymer composition obtained by the method of the present invention has a lower concentration of taspine than the concentration of taspin in the raw material, which is a milky juice.

In the composition obtained according to the methods proposed in the present invention, the proanthocyanidin polymer contains units representing leukoanthocyanidins. Leukanthocyanidins include, for example, monomeric flavonoids, which include catechins, epicatechins, gallocatechins, galloepicatechins, flavanols, flavonols and flavan-3,4-diols, leukocyanidins and anthocyanidins. In one embodiment, the proanthocyanidin polymer comprises polymers with flavonoid units from about 2 to about 30, from about 2 to about 15, from about 2 to about 11, or on average from about 7 to about 8 with a number average molecular weight of from about 2000 to about 3000 Da, or, for example, with a molecular weight of from about 1100 to about 2900 Da, or, for example, with a molecular weight of from about 1500 to about 3000 Da. The proanthocyanidin polymer can be dissolved in an aqueous solution, and the proanthocyanidin polymer can also be dissolved in other solutions and mixtures of aqueous and non-aqueous solutions.

In the composition obtained according to one embodiment of the invention, the proanthocyanidin polymer contains crofelemer (ϋδΑΝ). For more information on proanthocyanidin polymers and / or crofelemer, see, for example, SAZ, registration number 148465-456 and / or US patent No. 5211944 (Tstrscha).

Proanthocyanidin-based polymer compositions obtained by the method of the present invention can be analyzed by any methods known in the art. For example, proanthocyanidin polymers can be determined by absorption of ultraviolet radiation (lambda). Some proanthocyanidin monomers and polymers, for example, have broad peaks ranging from about 200 to about 300 nm, for example, in the range between about 190 and about 215 nm (e.g., about 205-210 nm) and between about 260 and about 295 nm (e.g., 275-280 nm). Fractions containing proanthocyanidin polymers can have an additional major UV absorption maximum in the range of about 400 to about 500 nm, between 425 and 475 nm, and about 460 nm.

Obtaining polymer compositions based on proanthocyanidin.

The present invention provides methods for producing a proanthocyanidin-based polymer composition having the degree of purity and concentration necessary for use in a therapeutically effective pharmaceutical composition. The starting material contains milky juice, for example, isolated from Crgoi crp. or Ca1orNu11it crr. Milky juice can be obtained by any method known in the art, for example, by cutting a croton tree and collecting milky juice that accumulates inside or protrudes from the incisions.

The method includes, for example, one or more of the following steps: production or preparation of a milky juice containing a proanthocyanidin polymer, keeping the milky juice (for example, obtained from Crgoi cr.) At a temperature below room temperature (for example, from about 0 to about 20 ° C., constantly or intermittently) for at least 48 hours (in the range between about 1 hour and about 30 days) to allow sedimentation of suspended particles; adjusting the pH of the milky juice to a value in the range of from about 6.5 to about 8.5; filtering milk juice to obtain a filtrate; adjusting the pH of the filtrate to a value in the range of about 3.5 to about 5.5; performing a first extraction of the filtrate using a first extraction phase (e.g., solid phase extraction) to obtain an eluate containing a proanthocyanidin polymer; performing a second extraction using a second extraction phase (for example, a solid phase) to obtain an eluate containing a proanthocyanidin polymer, and processing the eluate to obtain a polymer composition based on proanthocyanidin. The method may also include an additional, optional step, such as the implementation of an additional processing step, for example, an additional processing step between the first and second extraction.

In some embodiments, holding below room temperature includes one or more keeping the material below room temperature at all times, keeping the material at room temperature for most of the time, holding the material at room temperature intermittently, cyclically changing the temperature of the material, including temperature below room temperature. Aging may also include, for example, the time required to lower the temperature to below room temperature and the time to warm to room temperature or higher.

In one embodiment, the second extraction phase is different from the first extraction phase.

Alternatively, a method for producing a proanthocyanidin-based polymer composition may include preparing or providing a partially purified proanthocyanidin-based polymer composition; performing a first extraction of a partially purified proanthocyanidin-based polymer composition using a first extraction phase to obtain an eluate containing a proanthocyanidin polymer, and treating the eluate to obtain a purified proanthocyanidin-based composition, the first extraction being carried out using a solid-phase extraction resin.

In the invention, unless otherwise indicated, the term purified or pure means a composition that contains a polymer composition based on proanthocyanidin, practically free of impurities. For example, a pure proanthocyanidin-based polymer composition contains more than about 80 wt.% Proanthocyanidin-based polymer composition and less than about 20 wt.% Impurities, more preferably more than about 90 wt.% Proanthocyanidin-based polymer composition and less than about 10 wt.% impurities, even more preferably more than about 95 wt.% polymer composition based on proanthocyanidin and less than about 5 wt.% impurities. The degree of purity can be determined by methods known to the chemist or pharmacist. Preferably, the purity of the compounds is more than 99% (w / w), while compounds whose purity is more than 95, 90 or 85% can be used if necessary.

- 8 023814

Alternatively, along with or instead of solid-phase extraction, a method for producing a polymer composition based on proanthocyanidin may include a filtration step using a filter material (for example, bentonite, charcoal or one or more filter materials specified in the present invention), said milky juice or a partially purified proanthocyanidin-based polymer composition is mixed with a solvent such as alcohol (e.g. methanol), water, an aqueous / organic mixture, or under one of the above, and then washed or precipitated with another solvent, for example, ethyl acetate, methanol, ethanol, propanol, butanol, pentanol, hexanol, ethylene glycol, propylene glycol, dichloromethane, trichloromethane, tetrachloromethane, dichloroethane, diethyl ether, acetone, dimethyl ether, acetone their ether combinations, etc.

After completion of this method, a polymer composition based on proanthocyanidin contains a proanthocyanidin polymer and residual or trace amounts of, for example, water, other solvents used in the method, monomer units of the proanthocyanidin polymer, other components of natural origin that were part of the initial milky juice, etc. P. In one embodiment, the proanthocyanidin-based polymer composition of the present invention contains more than about 90% pure (e.g., determined by purity chromatography) proanthocyanidin polymer. Alternatively, the proanthocyanidin-based polymer composition prepared according to the process comprises more than about 95% pure proanthocyanidin polymer. In another embodiment, the proanthocyanidin polymer composition comprises a proanthocyanidin polymer and a residual amount of water. In another embodiment, the proanthocyanidin-based polymer composition mainly consists of a proanthocyanidin polymer and a residual amount of water. Preferably, the present invention comprises a proanthocyanidin polymer comprising more than 95% of a pure polymer composition based on proanthocyanidin, the amount of other components of which is less than 5%, and the amount of any other single component is less than 1.0%. The percentage by weight of the proanthocyanidin polymer can be obtained by methods known in the art, for example by HPLC analysis using a UV detector and measuring the area under the curve of the data obtained. The percentage by weight refers, for example, to the determined components in comparison with a standard sample and does not depend on the amount of moisture.

The lower limit of the purity determined, for example, by chromatography of compositions obtained according to the methods proposed in the present invention, can be an amount of proanthocyanidin polymer, for example, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99 %, or any amount in the range between the listed amounts. The purity of the compositions obtained according to the methods proposed in the present invention can reach 100% pure inclusive proanthocyanidin polymer, where an amount of 100% means that no additional components are found at a detectable level. Alternatively, the upper purity limit may be an amount of a proanthocyanidin polymer of 99.99, 99.9 and 99%, or any amount in the range between those listed.

The present invention provides methods for producing a proanthocyanidin-based polymer composition from milk juice containing a proanthocyanidin polymer. Milky juice is kept at a temperature below room temperature (for example, below 20 ° C) for a period of time sufficient to allow sedimentation of suspended particles (for example, from about 1 hour to about 30 days). A greater amount of time may be allowed to settle suspended particles, for example from about 30 days to about 4 years. In one embodiment, the milky juice may be infused at a temperature of about 0-15 ° C. for a period of time from 48 hours to 48 months, or at a temperature value or for a period of time within an appropriate range, for example, at 0-10 ° C, 2-8 ° C or 5 ° C for 13, 14 or 15 days. In one embodiment of the invention, the milk juice is sedimented for 14 days at a temperature of 2-8 ° C. Alternatively, any value in the above ranges may be applied.

In the present invention, the time sufficient for deposition is the time during which the settling of suspended particles on or near the bottom of the vessel containing the polymer will be ensured. The indicated time may be, for example, 1 hour or 30 days. Adequate time can also be a set average time needed to ensure a sufficient degree of subsidence.

After a sufficient time has elapsed for precipitation at a temperature below room temperature, the pH of the milky juice is adjusted to a value between about 6.5 and about 8.5. The pH can be adjusted using any pH adjusting agent known to one skilled in the art, for example, sodium carbonate, sodium bicarbonate, sodium hydroxide, calcium carbonate, and the like. In one embodiment of the invention, the pH is adjusted to about 8. After adjusting the pH, the milky juice is sedimented. For example, in one embodiment of the invention, the pH adjusted milk juice is cooled for 2 hours in order to ensure receipt and precipitation from the milk

- 9,023,814 solids juice or suspension. The degree of cooling can vary from a temperature below room temperature to the freezing temperature of milky juice. For example, the temperature may range from about 0 to about 15 ° C.

After adjusting the pH and the precipitation period, the reaction mixture was passed through filter material. Suitable filter materials contain diatomite (available as Ce1a1ot® or CeChe®). Other filter materials may contain cellulose mesh filter media, for example perlite, charcoal, bentonite, a glass filter element, sand, filter paper, Oo \\ e. \. salt or similar materials. At the end of the filtration, the solid residue on the filter material may be washed with additional aliquots of an aqueous solution or water. If desired, the filtration step may include centrifugation to assist in the filtration step.

In one embodiment of the invention, bentonite and / or one or more of the other filter materials listed are used either as a single filter material or in combination with one or more other filter materials in an aqueous, methanol, aqueous-organic mixture or other solvent system to reduce the level of protein, and / or taspine, and / or related substances, and / or impurities after filtering. Bentonite is a commercially available clay. For example, bentonite can be purchased from епΙΦποΙι SNepisan. Other filter materials may include cellulose mesh filter media, such as perlite, charcoal, bentonite, a glass filter element, sand, filter paper, Oo ^ ex, salt or similar materials. Not wanting to be associated with any theory, however, it is believed that at a certain pH, protein, and / or taspin, and / or related substances, and / or impurities that may be present in the starting material, which is milky juice, or partially purified polymer compositions based on proanthocyanidin can exist in the form of positively charged particles that interact with a negative charge on bentonite or other filter material, which leads to the removal of protein and / or taspine and / or related substances and / or and impurities together with bentonite or other filter material after filtration. In one embodiment of the invention, bentonite is added to an aqueous, or methanol, or aqueous-organic system containing a partially purified proanthocyanidin-based polymer composition and mixed, followed by filtration to remove bentonite and any materials or particulate particles associated with bentonite. The amount of protein removed in this way can be more than about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, and about 95% and about any integer value in the range between the indicated values compared to the amount of protein in the original milky juice. For example, the use of bentonite in the water system can help remove more than about 50% of the protein compared to the amount in milky juice. Bentonite or other filter material can be applied separately or in combination with other steps mentioned in this method, for example, solid phase extraction, precipitation of crystals from a solution, etc.

The resulting filtrate can be combined with water, which washed the filter material, and the combined aqueous layers can be treated to remove excess water and / or prepare the solution for further processing. Processing and / or concentration can be achieved in vacuo or, for example, by ultrafiltration (for example, using a semi-permeable membrane). In one embodiment, the concentration is carried out using ultrafiltration. Not wanting to be associated with any theory, it is believed that ultrafiltration provides the retention of suspended solids and dissolved substances with a high molecular weight on one side of a semipermeable membrane, while water and dissolved substances with a low molecular weight pass through the membrane. Corresponding semipermeable membranes may have a delay threshold of 500 Da, 1, 5, 10, 20, 30 kDa, 500 Da to about 750, 900 Da to about 5 kDa, or 1 to about 40 kDa. If desired, ultrafiltration can occur more than once, or more than one type of semi-permeable membrane can be applied simultaneously or sequentially with a portion being separated using a higher and lower permeability threshold. Using one or more types of semipermeable membranes, the set of chemical parameters (for example, the number and size of components of natural origin that were part of the initial milky juice, with the exception of the proanthocyanidin polymer) and the polydispersity index of the proanthocyanidin polymer (for example, the molecular weight distribution of the polymer) of the obtained composition can be changed . In one embodiment of the invention, the semi-permeable membrane allows the passage of dissolved low molecular weight substances, the molecular weight of which does not exceed 1 kDa. As an alternative, a semipermeable membrane will retain solutes with a molecular weight of more than 5 kDa.

After processing the filtrate and optionally washing, the pH of the resulting solution may be

- 10 023814 brought to a value in the range from 3.5 to 5.5 using, for example, a solution of a strong or weak acid containing hydrochloric acid, citric acid, etc. In one embodiment of the invention, the pH is adjusted to about 4. Alternatively, the pH can be adjusted before ultrafiltration by the method described above.

In one embodiment, after treatment with bentonite or another filter material, milk juice, or an aqueous solution, or a methanol solution, or an aqueous-organic system containing milk juice, is treated with ethyl acetate one or more times. Suitable solvents include, for example, methanol, ethanol, propanol, butanol, pentanol, hexanol, ethylene glycol, propylene glycol, ethyl acetate, dichloromethane, trichloromethane, carbon tetrachloride, dichloroethane, diethyl ether, acetone, dimethylformamide, dimethyl sulfoxide and combinations thereof . Not wishing to be bound by any theory, it is believed, however, that ethyl acetate removes low molecular weight related substances, while predominantly leaving the desired proanthocyanidin polymers. Low molecular weight related substances may include, without limitation, subsequently leachable materials, possibly low molecular weight phenolic compounds. Processing with ethyl acetate may occur one or more times. For example, two, three, four, five, six, seven, eight, nine, ten or more rinses with ethyl acetate can be carried out. The pH can be adjusted to a value in the range of from about 6.5 to about 8.5, for example, using 0.5N. ΝαΟΗ solution. In one embodiment, six washes with ethyl acetate are performed. Since, after washing with ethyl acetate, an aqueous, or methanol, or water-organic layer exists, the desired proanthocyanidin polymer may remain in the aqueous layer and, thus, may be separated from the ethyl acetate layer for purification. Alternatively, ethyl acetate can be considered as a co-solvent, which when added by itself or with other precipitating agents, such as salt, causes precipitation from the solution. In other words, since there is a lack of water, or methanol, or an aqueous-organic system to retain the proanthocyanidin-based polymer composition in the solution due to the addition of ethyl acetate, a precipitate may form which can be separated from the liquid layers by filtration. Reversible aqueous extraction of ethyl acetate used for washing can also be carried out. In one embodiment of the invention, the obtained proanthocyanidin-based polymer composition has an average molecular weight in the range of from about 2000 to about 3000 Da, or, for example, a molecular weight in the range of from about 1100 to about 2900 Da, or, for example, a molecular weight in the range from about 1500 to about 3000 Yes and a polydispersity in the range between about 1.2 and about 1.8, or, for example, between about 0.9 and about 1.2, or from about 0.5 to about 1.5, or from about 0.8 to about 1.3.

In one embodiment of the invention, both bentonite and ethyl acetate are treated. In an alternative embodiment, treatment with bentonite and ethyl acetate occurs before, after, or instead of any solid phase extraction steps. Preferably, the amount of the proanthocyanidin-based polymer composition isolated after treatment with bentonite and / or ethyl acetate is more than 30 wt.% Compared to the starting material. Alternatively, the amount of the product may be more than 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50% or more .

Solid-phase extraction of a pH-adjusted solution provides additional purification of the proanthocyanidin polymer. Solid phase extraction can be carried out using various solid phases, such as cross-linked polystyrene, modified agarose or modified dextran. The type of solid phase is not limited and may include resins such as ion exchange resins, adsorption resins, separation resins, size exclusion chromatography resins, mixed resin and mixtures of more than one type of resin. In one embodiment of the invention, solid phase extraction is carried out using a resin with some ion exchange properties, such as carboxymethyl modified agarose. Alternatively, the solid phase extraction resin may be a modified polysaccharide, such as hydroxypropylated cross-linked dextran. Commercially available carriers for solid phase extraction include CM-§peryago8e®, which is a carboxymethyl-modified agarose, and §erbabehH20®, which is a hydroxy-propylated cross-linked dextran. Solid phase extraction can be carried out in batch or continuous mode. In one embodiment of the invention, solid phase extraction is carried out batchwise to increase efficiency. After extraction, the solid phase is recovered to remove the proanthocyanidin polymer from the solid phase. The recovery can be carried out using any solvent or mixture of solvents that remove proanthocyanidin from the solid phase. For example, extraction solvents contain water, acetone, methanol, ethanol, glycol, and mixtures thereof. In one embodiment of the invention, the extraction is carried out using water and mixtures of water and acetone.

In the above methods, the eluate may be subjected to additional solid phase cycles

- 11,023,814 extraction using a solid phase different from the solid phase used previously. The type of solid phase is not limited and may include, for example, resins such as ion exchange resins, adsorption resins, separation resins, size exclusion chromatography resins, mixed resin and mixtures of more than one type of resin. For example, in the case where CM-Speryago5e® is used to carry out the first solid-phase extraction, the second solid-phase extraction can be carried out using Zerjeh L-20® as a solid support, or vice versa.

After performing solid phase extraction or, alternatively, after applying a filter material such as bentonite, an eluate containing a proanthocyanidin polymer may be mixed with a solvent selected from one or more solvents including methanol, ethanol, propanol, butanol, pentanol, hexanol , ethylene, propylene glycol, ethyl acetate, dichloromethane, trichloromethane, carbon tetrachloride, dichloroethane, diethyl ether, acetone, dimethylformamide, dimethyl sulfoxide, ether, combinations thereof and the like. Not wanting to be associated with any particular theory, mixing with an additional solvent may be useful to provide additional deposition and / or additional washing of the layer containing the proanthocyanidin polymer. In one embodiment, the solvent for treating the aqueous concentrate is butanol. After optional mixing with an additional solvent, the proanthocyanidin polymer-containing layer may be treated by ultrafiltration, evaporation with or without heating, evaporation with or without vacuum, freeze-drying, spray drying and the like, including combinations of processing technologies, to obtain a polymer composition based on proanthocyanidin.

According to the methods proposed in the present invention, proanthocyanidin-based polymer compositions have concentrations and purities suitable for use in pharmaceutical compositions. The lower limit of the purity determined by chromatography of the compositions obtained according to the methods of the present invention may be an amount of proanthocyanidin polymer of about 80, about 90, about 91, about 92, about 93, about 94, about 95, about 96, about 97, about 98, or about 99%, or any amount in the range between the listed amounts. The purity determined by chromatography of the compositions obtained according to the methods proposed in the present invention can reach 100% inclusive, where an amount of 100% means that there are no additional components at a detectable level. Alternatively, the upper limit of purity determined by chromatography may be about 99.99, about 99.9, and about 99%, or any amount in the range between those listed. For example, the amount of taspine present in the original milky juice can be reduced by the methods of the present invention. The level of taspine, the purity of which is determined by chromatography, in the polymer composition based on proanthocyanidin obtained according to the methods proposed in the present invention, can vary from 1% to amounts below the detectable level. For example, the upper limit of the amount of taspine, the purity of which is determined by chromatography, in a polymer composition based on proanthocyanidin can be about 1.0, about 0.9, about 0.8, about 0.7, about 0.6, about 0.5 %, about 0.4, about 0.3, about 0.2, about 0.1, or about 0.05%, or any amount in the range between the listed amounts or below, up to the detection level. In one embodiment of the invention, in the polymer composition based on proanthocyanidin, the level of taspine, the purity of which is determined by chromatography, is below about 0.2% (for example, 2000 ppm). The proanthocyanidin-based polymer composition obtained according to the methods of the present invention preferably contains an amount of taspin, the purity of which is determined by chromatography, in the range between about 0.1 and about 0% (for example, no detectable amount of taspin).

In one embodiment of the invention, the polydispersity index may vary from about 1.2 to 1.8. Depending on the desired pharmaceutical form, a narrow range of molecular weights can be chosen to achieve a low polydispersity, or a wider range of molecular weights, or various higher or lower thresholds, can be selected. Polydispersity is determined not only by the indicated ultrafiltration step, but may also depend on other parameters of the method.

As an alternative to the above method, the methods proposed in the present invention also include obtaining a purified polymer composition based on proanthocyanidin in the degree of purity and concentration necessary for inclusion in a therapeutically effective pharmaceutical composition, including obtaining a partially purified polymer composition based on proanthocyanidin, performing extraction ( for example, solid phase extraction) of a partially purified proanthocyanidin-based polymer composition to give elu an ata containing a proanthocyanidin polymer and treating the eluate to obtain a purified proanthocyanidin composition. In one embodiment of the invention, the extraction is carried out using

- 12 023814 resins for solid phase extraction as a solid phase. In one embodiment of the invention, the partially purified proanthocyanidin-based polymer composition is partially purified by a method involving the production of milky juice from CgOup BPP. and performing solid phase extraction using a resin for solid phase extraction, which is not hydroxypropylated cross-linked dextran. In one embodiment of the invention, the solid phase extraction resin used to partially purify the starting material is a carboxymethyl modified agarose resin. A partially purified proanthocyanidin-based polymer composition may contain, for example, from about 35 to about 90% of the proanthocyanidin polymer, the purity of which is determined by chromatography. In the range of from about 35 to about 90%, the partially purified proanthocyanidin-based polymer composition may contain any sub-range, for example 40-80, 45-75, 50-70, 55-65, or about 60%. Alternatively, the partially purified proanthocyanidin-based polymer composition may contain any numerical amount of the proanthocyanidin polymer in the range of 35-90%, for example 36, 37, 38%, and the like. up to 87, 88, 89 and 90% or any value between them. In one embodiment, the partially purified proanthocyanidin-based polymer composition may be 8B-300. In addition, the partially purified proanthocyanidin-based polymer composition may have a taspine level of about 0.2% or higher, for example about 0.2, about 0.3, about 0.4, about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1.0% or higher, or any amount in the range between the listed level values up to the natural level in milky juice.

Solid phase extraction can be performed using one or more resins for solid phase extraction. For example, such resins may include modified polysaccharides or similar materials such as carboxymethyl modified agarose. Modified polysaccharides may include hydroxypropylated cross-linked dextran. Commercially available carriers for solid phase extraction include CM-§perNato8e®, which is a carboxymethyl-modified agarose, and §ERNaye \ LH20®, which is a hydroxy-propylated cross-linked dextran.

After performing solid phase extraction, the eluate containing the proanthocyanidin polymer can be mixed with a solvent selected from one or more solvents including methanol, ethanol, propanol, butanol, pentanol, hexanol, ethylene glycol, propylene glycol, ethyl acetate, dichloromethane, trichloromethane, tetrachloride, tetra diethyl ether, acetone, dimethylformamide, dimethyl sulfoxide, ether, mixtures thereof, etc. In one embodiment, the solvent for washing the aqueous concentrate is butanol. After optional mixing with an additional solvent, the layer containing the proanthocyanidin polymer can be processed by ultrafiltration, evaporation with or without heating, evaporation with or without vacuum, freeze drying, spray drying, etc., including combinations of processing technologies, to obtain the finished polymer composition based on proanthocyanidin.

In the method for producing proanthocyanidin, organic solvents are selected from one or more solvents, including alcohols, ketones, esters and ethers. Ketones are selected, for example, from methyl ethyl ketone and isobutyl ketone. Esters are selected, for example, from ethyl acetate, methyl acetate, propyl acetate and butyl acetate. Ethers are selected, for example, from diethyl ether, methyl ether, dimethyl ether, etc. Organic solvents can also be mixtures of alcohols, ketones, esters and ethers. For example, the mixture may be a combination of alcohol and esters such as methanol and ethyl acetate.

The chromatographic purification system contains, for example, a separate column or an installation from columns selected, for example, from CM-ePatOnate8Pa1 Iote CoItp and Sperie / LH-20. When using multiple columns, they can be used together, one after another or separately. Eluents are selected, for example, from aqueous solvents such as water and solvents miscible with water.

In another embodiment, a method for producing a proanthocyanidin-based polymer composition may include purification methods and may include creating partially purified milky juice dissolved in alcohols (e.g. methanol) and extracting the dissolved compound with alcohol, followed by treatment using a filter (e.g. charcoal ) and filtration through another filter layer, for example a bentonite layer. The filtrate can then be treated with acetate (e.g., ethyl acetate), followed by passing through a filter (e.g., HUN-Papp) to give proanthocyanidin without taspine impurities, for example, where the amount of taspin is below the detectable level.

Therapeutic applications.

The invention describes the use of compositions obtained according to the production methods set forth in the invention for the treatment and / or prevention of one or more symptoms of irritable bowel syndrome with predominant diarrhea (ROS) in warm-blooded animals, including

- 13 023814 male and female people, these symptoms include, but are not limited to, pain, abdominal discomfort, diarrhea, the presence of an urgent urination, violation of the frequency and consistency of the stool. The methods described herein typically include administering to a subject in need of treatment for ROS, a proanthocyanidin-based polymer composition. In one embodiment of the invention, the proanthocyanidin-based polymer composition is orally administered and is not absorbed systemically. In one embodiment, the subject comprises an animal. In one embodiment, the subject comprises a primate. In another embodiment, the subject includes a human. In another embodiment, the subject includes a woman and / or a man.

In one embodiment of the invention, methods of treating pain and diarrhea caused by RDS are provided, comprising administering to a subject in need of such treatment a proanthocyanidin polymer composition in an amount effective to treat pain and diarrhea caused by RDS. In another embodiment, the present invention provides methods for treating gastrointestinal discomfort and diarrhea caused by RDS, comprising administering to a subject in need of such treatment a polymer composition based on proanthocyanidin in an amount effective to treat gastrointestinal discomfort and diarrhea caused by RDS. In some embodiments of the invention, a certain amount of the proanthocyanidin-based polymer composition is administered together with an analgesic and / or anti-inflammatory compound, for example, an inhibitor of COX-2 and preferably an inhibitor of COX-2 rather than COX-1.

In other embodiments, a proanthocyanidin-based polymer composition is administered either simultaneously, before, during, or after the administration of another composition.

In typical embodiments of the invention, the frequency of the stool is reduced by at least 10, 20, 30, 40 50 60 70 80% or more compared with the frequency of the stool of the subject prior to treatment using a polymer composition based on proanthocyanidin. In other embodiments of the invention, the frequency of the stool is reduced by at least one bowel movement per day compared to the amount before treatment with a polymer composition based on proanthocyanidin. In other embodiments of the invention, the stool consistency is compacted, for example, the amount of water in the stool is reduced by at least 10, 20, 25, 30, 40, 50, 60, 70, 80% or more compared with the consistency before treatment. In other embodiments of the invention, the urgent urination to urinate in a subject is reduced by at least 10, 20, 30, 40, 50, 60, 70, or 80% more than the urge to initiate treatment using a polymer composition based on proanthocyanidin.

In another embodiment of the invention, there is provided a method for treating diarrhea due to RDS, which comprises orally administering to a patient in need of such treatment a proanthocyanidin-based polymer composition, for example enteric-coated crofelemer (CL 148465-45-6), in an amount effective for the treatment of diarrhea due to ROS, while the indicated amount is in the range between about 50 mg per day and about 750 mg per day, and crofelemer is obtained by the method proposed in the present invention. In another embodiment, the present invention provides a method for treating a violation of stool frequency, violation of stool consistency, or having an urgent urination caused by an RDS, comprising orally administering to a patient in need of such treatment, enteric coated crofelemer in an amount effective to treat a frequency violation stool, a violation of the consistency of the stool or the presence of an urgent urination caused by ROS, while the indicated amount is in the range between about 5 0 mg per day and about 750 mg per day, and crofelemer obtained by the method proposed in the present invention.

The pharmaceutical forms described in this application can also be used to treat irritable bowel syndrome with predominant diarrhea (RDS) in non-human animals, in particular farm animals such as, but not limited to, cattle, pigs, sheep and rams, birds (such as chickens) and horses and other domestic animals such as dogs and cats. In particular, the pharmaceutical forms described herein can be used to treat non-human RDS diseases in animals, in particular beef and dairy animals such as cattle, sheep and pigs, by incorporating a polymer composition based on proanthocyanidin or their pharmaceutical compositions in animal feed.

In determining whether a subject with IBS suffers from predominant diarrhea, any method accepted in the art can be used to diagnose the subject, including, but not limited to, Rome criteria II for diagnosing irritable bowel syndrome (Thotrkop et al., 1999, Oi! 45 (ext. II): Ш43-1147). In brief, diagnostic Rome II criteria state that over the past 12 months for at least 12 weeks, not necessarily consecutively, discomfort or abdominal pain, the subject experiences two of the following three symptoms: (1) relief from bowel movements, (2) onset associated with a change in stool frequency; and (3) the onset is associated with a change in the shape (appearance) of the chair. The following symptoms in total confirm the diagnosis of IBS: (ί) violation of stool frequency, for example, more than 3 times a day; (i) impaired stool shape, for example liquid / watery stools; and (ίίί) the presence of an urgent urination (urge to defecate).

The degree of pain and discomfort can be measured by any method known in the art. For example, a pain or discomfort scale can be applied, with the patient evaluating the level of pain or discomfort on a scale from 0 to 5, where 0 corresponds to the absence of pain or discomfort, and 5 corresponds to the highest level of pain or discomfort. In some embodiments, the relief of pain or discomfort is determined by lowering the average level of pain or discomfort and / or increasing the number of days without pain or discomfort. In some embodiments, the number of days without pain or discomfort is increased by at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90% or more compared to the number before treatment with a polymer composition based on proanthocyanidin. In other embodiments, the level of pain or discomfort is reduced by at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.75, 1.0, 1.5, or 2.0, or more units compared to the level before treatment.

In another embodiment, the present invention provides methods of treating irritable bowel syndrome with predominant constipation (SRKZ). The methods include administering to a subject in need of treatment for SRH, a proanthocyanidin polymer composition. In one embodiment of the invention, the proanthocyanidin-based polymer composition is orally administered and is not absorbed systemically. In one embodiment, the patient is a female person. In another embodiment, the present invention provides methods for treating pain or any other symptoms or a combination of symptoms caused by SRH, comprising orally administering to the patient in need of such treatment, enteric-coated crofelemer (CA§ 148465-45-6) in an amount effective for the treatment of pain due to SRH, wherein said amount is in the range between about 500 mg per day and about 3 g per day, and crofelemer is produced by the method of the present invention.

In another embodiment of the invention, methods of treating secretory diarrhea using the composition of the present invention are provided. Pharmaceutical compositions are used to treat diarrhea of travelers and nonspecific diarrhea. In another embodiment of the present invention, pharmaceutical compositions prepared according to the methods provided herein are used to treat secretory diarrhea caused by viral infections, such as diarrhea, which are caused by an infection caused by human immunodeficiency virus (HIV) and acquired immunodeficiency syndrome (AIDS) ) and rotavirus infection. Almost all AIDS patients suffer from diarrhea at some point in the course of the disease, and 30% of AIDS patients suffer from chronic diarrhea. Diarrhea, which is caused by AIDS and HIV infection, has been called HIV-associated chronic diarrhea. The indicated diarrheal component of the disease caused by HIV is believed to be due to secondary infections, which include protozoal pathogens, for example, SgurYukropShit krr., HIV itself or therapy used to control viral load. Pharmaceutical compositions are used to treat the above secretory diarrhea associated with HIV and AIDS in adults and children. In addition, rotavirus infection is a significant cause of diarrhea, in particular in infants and young children in developing countries, for which the pharmaceutical compositions of the present invention are used.

This application describes the use of compositions obtained according to the production methods set forth in this application for the treatment and / or prevention of colon cancer in animals, including males and females. In particular, the present invention provides methods for treating or preventing colon cancer, comprising administering to a patient in need of such prophylaxis, a certain amount of a proanthocyanidin-based polymer composition obtained by the methods of the present invention. For example, a patient in need of such treatment or prophylaxis includes those patients who have been diagnosed with colon polyps or those who have been diagnosed with colon cancer (for example, colon cancer at any of stages 0, I, II, III, or IV), or those patients who have previously been treated for colon cancer (for example, to prevent relapse), or those patients who have a family or hereditary predisposition to colon cancer, or patients living in areas in which the incidence of cancer is thick gut above the average norm. Other patients requiring such treatment or prophylaxis are those who underwent a colon biopsy showing precancerous changes.

The methods of the present invention comprise administering to a subject in need of treatment of colon cancer a proanthocyanidin polymer composition. In one embodiment of the invention, the proanthocyanidin-based polymer composition is orally administered and is not absorbed systemically. In one embodiment, the subject comprises an animal. In one embodiment, the subject comprises a primate. In another embodiment, the subject includes a human. In another embodiment, the subject includes a woman and / or a man.

In some embodiments of the invention, for the treatment of colon cancer, a certain amount of the proanthocyanidin-based polymer composition is administered together with an anesthetic and / or anti-inflammatory compound, for example, an inhibitor of COX-2 and preferably an inhibitor of COX-2 rather than COX-1.

In determining whether a subject suffers from colon cancer or whether there is an increased risk of developing colon cancer, any method adopted in the art can be applied.

In some embodiments of the invention, the effectiveness of the treatment of colon cancer is determined by reducing the size of the colon tumor. In some embodiments of the invention, the size of the colon cancer is reduced by at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90% or more compared to up to initiation of treatment using a polymer composition based on proanthocyanidin. In other embodiments of the invention, the effectiveness of the treatment of colon cancer is determined by the absence of an increase in the growth of a colon cancer.

In one embodiment of the invention, the patient is a human, for example, a male or female. In another embodiment of the invention, there is provided a method for treating colon cancer, comprising orally administering to the patient in need of such treatment, enteric-coated crofelemer (SAB 148465-45-6) in an amount effective for treating colon cancer, wherein said amount is in the range between about 500 mg per day and about 3 g per day, and crofelemer obtained by the method proposed in the invention.

In some embodiments of the invention, proanthocyanidin-based polymer compositions may be administered as adjuvant therapy with other known therapies for treating or preventing colon cancer. For example, the compositions may be administered before, simultaneously with or after surgery, radiation therapy, chemotherapy or biological therapy (biological therapy includes immunological therapy using engineered antibodies such as ΕΚΒΙΤυΧ®, ΑνΑδΤΙΝ®, or therapy to enhance the immune response to cancer).

Pharmaceutical doses.

When applying the forms and methods proposed in the present invention for the treatment of pathological conditions, diseases and disorders, the effective dosage range of the pharmaceutical forms of the proanthocyanidin-based polymer composition for oral administration is in the range from 0.1 to 100 mg / kg per day, for example from about 0.1 to about 40 mg / kg per day; from about 0.1 to about 25 mg / kg per day; or from about 0.1 to about 10 mg / kg per day. It should be noted that the appropriate dose will depend on the type and severity of the condition or disease. It has been found that human subjects can tolerate at least 2 g of a polymer composition based on proanthocyanidin per day (25-30 mg / kg / day) for a period of up to 27 days.

In another embodiment of the invention, the pharmaceutical composition comprises a proanthocyanidin-based polymer composition obtained from Cίττοίοη crr., The dose of which does not exceed 750 mg per day, preferably less than 250 mg / day. In one embodiment of the invention, the proanthocyanidin-based polymer composition is crofelemer (SAB 148465-45-6).

In other embodiments, crofelemer is orally administered in an enteric-protected form (enteric-coated) in a total amount of not more than about 750 mg / day. In the present invention, approximately means within the margin of error. In specific embodiments of the invention, enteric-coated crofelemer is orally administered to the subject in an amount of about 50 to 750 mg / day. In another embodiment, the enteric-coated crofelemer is orally administered to the subject in a total amount of not more than about 500 mg / day. In specific embodiments, an enteric-coated crofelemer is orally administered to the subject in an amount of about 50 to 500 mg / day. In other embodiments, the enteric-coated crofelemer is orally administered to a subject in an amount of not more than about 700, about 650, about 600, about 550, about 500, about 450, about 400, about 350, about 300, about 250, about 200, about 150, or about 100 mg / day. In another embodiment, the enteric-coated crofelemer is orally administered to the subject in an amount of about 100 to 750 mg / day. In other embodiments of the invention, enteric-coated crofelemer is orally administered to a subject in an amount of from about 125 to about 500 mg / day, from about 250 to about 500 mg / day, from about 250 to about 450 mg / day, from about 250 to about 400 mg / day, about 250 to about 350 mg / day, or about 250 to about 300 mg / day. In other specific embodiments of the invention, the total dose of enteric-coated crofelemer orally administered to the subject one, two or three times a day is about 50, about 55, about 60, about 65, about 70, about 75, about 80, approximately 85, approximately 90, approximately 95, approximately 100, approximately 105, approximately 110, approximately 115, approximately 120, approximately 125, approximately 130, approximately 135, approximately 140, approximately 145, approximately 150, approximately 155, approximately 160, approximately 165 , about 170, about 175, about 180, about 185, about 190, about 195,

- 16,223,814 approximately 200, approximately 205, approximately 210, approximately 215, approximately 220, approximately 225, approximately 230, approximately 235, approximately 240, approximately 245, approximately 250, approximately 255, approximately 260, approximately 265, approximately 270, approximately 275 approximately 280, approximately 285, approximately 290, approximately 295, approximately 300, approximately 305, approximately 310, approximately 315, approximately 320, approximately 325, approximately 330, approximately 335, approximately 340, approximately 345, approximately 350, approximately 355, approximately 360, approximately 365, approximately 370, approximately 375, approximately 380, approximately 385, approximately 390, approximately 395, approximately 400, approximately 405, approximately 410, approximately 415, approximately 420 approximately 425, approximately 430, approximately 435, approximately 440, approximately 445, approximately 450, approximately 455, approximately 460, approximately 465, approximately 470, approximately 475, approximately 480, approximately 485, approximately 490, approximately 495, or approximately 500 mg day.

According to the methods for treating farm animals with ROSA proposed in the present invention, pharmaceutical compositions containing a proanthocyanidin-based polymer composition are administered to a subject in a total amount that is bioequivalent to not more than 750 mg / day of an orally administered enteric-coated proanthocyanidin polymer composition. In specific embodiments of the invention, pharmaceutical compositions comprising a proanthocyanidin-based polymer composition are administered to a subject in an amount of between about 50 and about 250 mg / day. In particular embodiments, the amount may be in the range of from about 50 to about 450 mg per day, or from about 50 to about 400 mg per day, or from about 50 to about 350 mg per day, or from about 50 to about 300 mg per day, or from about 50 to about 250 mg per day, or from about 50 to about 200 mg per day, or from about 50 to about 150 mg per day, or from about 50 to about 100 mg per day.

Proanthocyanidin-based polymer compositions can be combined in a pharmaceutical dose to administer a single or fractional dose one, two, three or four times a day. In a particular embodiment of the invention, the pharmaceutical dose is administered twice daily. In another embodiment, the pharmaceutical dose is administered twice daily for at least two consecutive days. In another embodiment, the pharmaceutical dose is administered at least for a period of time selected from the group consisting of 24, 48, 72, 96 hours, one week, two weeks, one month, two months and three months. In some embodiments of the invention, the pharmaceutical dose is taken for an indefinite period.

Pharmaceutical Forms

In another embodiment, a pharmaceutical form is provided comprising a proanthocyanidin-based polymer composition.

Methods for administering a polymer composition based on the proanthocyanidin of the present invention include, but are not limited to, parenteral administration (e.g., intradermal, intramuscular, intraperitoneal, intravenous and subcutaneous) and mucosal administration (e.g., intranasal and oral routes). In a specific embodiment, the proanthocyanidin-based polymer composition is administered intramuscularly, intravenously or subcutaneously. A polymer composition based on proanthocyanidin can be introduced in any convenient way, for example, by infusion or bolus injection, by absorption through the epithelial or skin-mucous lining (for example, the mucous membrane of the oral cavity, rectum and intestines, etc.) and can be introduced together with other biologically active agents. The introduction may be general or local. Preferably, the proanthocyanidin-based polymer composition is administered orally.

In one embodiment, the crofelemer comprises an enteric coating to protect against degradation by the acidic environment of the stomach and / or from interaction with proteins, such as pepsin, present in the stomach, for example, an enteric-soluble protected form. In a specific embodiment, crofelemer is used in the form of tablets. In yet another specific embodiment of the invention, the tablets are enteric coated, for example ΕυΌΚΛΟΙΤ®. In one embodiment, the crofelemer is prepared as an enteric-coated bead or granules in an enteric-coated capsule. In another embodiment, crofelemer is prepared in the form of a composition with a delayed release, for example, Meскsk ск, Αίζα ΘΚΘ δ, a wax matrix (the release is delayed mainly until the form passes from the stomach to the intestines).

In some embodiments, a proanthocyanidin-based polymer composition is prepared using a compound or compounds that neutralize stomach acid. Alternatively, a pharmaceutical composition containing inhibitor molecules is administered either simultaneously with, or after, or after administration of a pharmaceutical composition that neutralizes gastric acid. Compounds such as antacids that are used to neutralize stomach acid include, for example, aluminum carbonate, aluminum hydroxide, bismuth subnitrate, bismuth subsalicylate, calcium carbonate, sodium dihydroxyaluminium carbonate, magaldrate, magnesium carbonate, magnesium hydroxide, magnesium oxide, and mixtures thereof. Compounds that are capable of reducing the excretion of gastric acid and / or are capable of decreasing the acidity of the gastric fluid are known in the art and include, for example, antacids (aluminum hydroxide, aluminum carbonate, aluminum glycinate, magnesium oxide, magnesium hydroxide, magnesium carbonate, calcium carbonate, bicarbonate sodium)

- 17 023814 blockers of the secretion of gastric acid (cimetidine (Tadate! ™), famotidine (Mu1ai1a ™, ReryL ™), nizatidine (Αχίά ™), ranitidine (Ζ; · ιηΙ; κ ™), omeprazole (2.deg10 ™)) and a combination of any of the above compounds. In general, any drug that is approved for sale by an appropriate government agency and capable of reducing the production of stomach acid and / or acidity of the stomach fluid can be administered in combination with a proanthocyanidin-based polymer composition such as crofelemer.

In other embodiments of the invention, the proanthocyanidin-based polymer compositions of the present invention are administered with other compounds that are used to treat diarrhea, pain, or cancer. These compounds include, but are not limited to, COX-2 inhibitors, such as 5-ACA, sulfasalazine, mesalamine, ΑΡΑΖΑ, as well as other commercially available COX-2 inhibitors, such as celecoxib and rofecoxib. Compounds for treating cancer include synthetic and biological drugs that specifically target cancer cells. Preferably, said compounds are not absorbed systemically or are modified so as not to be absorbed systemically.

In a particular embodiment of the invention in which crofelemer does not have an enteric coating, crofelemer is prepared using one or more compounds that are capable of decreasing the secretion of gastric acid and / or are capable of decreasing the acidity of the gastric fluid. Preferably, the dose of crofelemer in said form is a dose that is bioequivalent to an oral dose of enteric-coated crofelemer of about 50 to about 750 mg per day. In a typical embodiment, the crofelemer is formulated as a controlled release (delayed release) composition.

In other embodiments, the proanthocyanidin-based polymer composition may be administered in combination with painkillers or anti-inflammatory agents. In one embodiment of the invention, the analgesic or anti-inflammatory agent is prepared or modified in such a way that said agent is generally not absorbed systemically, for example, only 20, 15, 10, 5, 4, 3, 2, 1 or 0.5 are absorbed. % of the prescribed dose.

The proanthocyanidin polymer composition of the present invention may be provided in any therapeutically acceptable pharmaceutical form. The pharmaceutical composition can be prepared for oral administration, for example, but not limited to, in the form of drug powders, crystals, granules, microparticles (which include particles of the order of several micrometers, such as microspheres and microcapsules), particles (which include particles of the order of a few millimeters), balls, microspheres, pellets, pills, microtablets, compressed tablets or trituration tablets, molded tablets or trituration tablets and in capsules, which s are solid or soft and contain the composition as a powder, particle, bead, solution or suspension. The pharmaceutical composition can also be prepared for oral administration in the form of a solution or suspension in an aqueous solution, in the form of a liquid included in a gel capsule or in any other form suitable for administration, or for rectal administration in the form of a suppository, enema or in any other convenient form. Inhibitor molecules or anticancer agents can also be provided as a controlled release system (see, for example, Laideg, 1990, Zaepse 249: 1527-1533).

The pharmaceutical composition may also include any kind of pharmaceutically acceptable excipients, additives or media. For example, diluents or fillers such as dextrates, dicalcium phosphate, calcium sulfate, lactose, cellulose, kaolin, mannitol, sodium chloride, dry starch, sorbitol, sucrose, inositol, icing sugar, bentonite, microcrystalline cellulose or hydroxypropyl methylcellulose can be added to form for increasing the size of the composition. In addition, binders such as, but not limited to, starch, gelatin, sucrose, glucose, dextrose, molasses, lactose, gum arabic, sodium alginate, Irish moss extract, panwar gum, ghatti gum, seed husk glue isapgol, carboxymethyl cellulose, methyl cellulose, polyvinylpyrrolidone, ueedite and starch arabogalactan, polyethylene glycol, ethyl cellulose and waxes can be added to the form to improve the bonding properties. In addition to the form, lubricants may be added, such as, but not limited to, talc, magnesium stearate, calcium stearate, stearic acid, hydrogenated vegetable oils, polyethylene glycol, sodium benzoate, sodium acetate, sodium chloride, leucine, carbovax, sodium lauryl sulfate and magnesium lauryl sulfate. In addition, glidants, such as, but not limited to, colloidal silicon dioxide or talc, can be added to improve the flow properties of the powder form. Finally, disintegrants such as, but not limited to starches, clays, cellulose, algins, gums, cross-linked polymers (e.g. croscarmellose, crospovidone and sodium starch glycolate), ueedite, methyl cellulose, agar, bentonite, cellulose and wood foods, natural sponge, cation exchange resins, alginic acid, guar gum, citrus pulp, carboxymethyl cellulose or sodium lauryl sulfate with starch can also be added to facilitate intestinal disruption.

- 18 023814

In one aspect of the present invention, a proanthocyanidin-based polymer composition prepared according to the methods of the present invention is formulated for oral administration. In other aspects of the invention, a pharmaceutical dosage form is formulated to protect a proanthocyanidin-based polymer composition from degradation by the acidic environment of the stomach and from interaction with proteins, such as pepsin, present in the stomach. Thus, in a preferred aspect of the invention, said form comprises an enteric coating. For example, an enteric-coated form is enteric-coated tablets, balls or granules that optionally contain a lubricant, such as, but not limited to, magnesium stearate. Enteric-coated forms include enteric-coated capsules in a capsule, enteric-coated microspheres in a capsule, enteric-coated microspheres, provided as a suspension or mixed with food, these suspensions are particularly suitable for administration to children, and enteric-coated compressed tablets. The capsule may be a hard gelatin capsule or a cellulose capsule. In particular, the pharmaceutical composition is prepared in the form of an enteric-coated capsule. In one specific aspect of the invention, the proanthocyanidin-based polymer composition is administered in the form of tablets that are filled with microcrystalline cellulose.

In one embodiment of the invention, the proanthocyanidin-based polymer composition is directly compressed, that is, the proanthocyanidin-based polymer composition with or without any excipients can be compressed into a tablet or other pharmaceutical form, which has a pharmaceutically acceptable strength and abrasion. Preferably, the directly compressible pharmaceutical composition can be compressed into tablets with a strength of more than 4 kp (kilopond), preferably with a strength of 8 to 14 kp, more preferably with a strength of 10 to 13 kp. The directly compressible composition can be compressed into a tablet, the abrasion of which is not more than 1% weight loss, preferably less than 0.8% weight loss, more preferably less than 0.5% weight loss.

In one embodiment of the invention, directly compressible forms consist of 99.93% of a polymer composition based on proanthocyanidin and 0.07% of magnesium stearate and are coated with a copolymer with methacrylic acid. In another embodiment of the invention, the pharmaceutical form comprises a directly compressible polymer composition based on proanthocyanidin without excipients, additives or media except enteric coating; however, the form may contain a lubricant, such as, but not limited to, magnesium stearate. Preferably, the form of the directly compressible proanthocyanidin-based polymer composition is prepared as a tablet of pharmaceutically acceptable strength (more than 4 kp, preferably 8-14 kp and more preferably 10-13 kp) and abrasion (not more than 1% weight loss, preferably less than 0.8% weight loss, and more preferably less than 0.5% weight loss).

In another embodiment, the proanthocyanidin-based polymer composition is enteric coated. Enteric coatings are coatings that remain intact in the stomach but dissolve and release the contents of the dosage form as soon as the small intestine is reached. A large number of intestinal coatings are prepared using components containing such acid groups that, at very low pH values in the stomach, for example at pH values from 1.5 to 2.5, do not decompose into ions, and the coating remains in an undissociated, insoluble form. At higher pH values, such as in the intestinal environment, the enteric coating turns into an ionized form that can be dissolved to release the inhibitor molecules. Other types of enteric coatings remain intact until they are decomposed by enzymes in the small intestine, and the third types of coatings disintegrate after exposure to moisture, such that the coatings remain intact until they enter the small intestine.

Polymers that are used to form enteric coatings include, but are not limited to, shellac, starch and phthalates, amylose acetate, styrene-maleic acid copolymers, cellulose acetate succinate, cellulose acetate phthalate (CAF), polyvinyl acetate phthalate (PVAF) hydroxypropyl methyl cellulose phthalate (brands HP-50 and HP-55), ethyl cellulose, fats, butyl stearate and copolymers of methacrylic acid and methacrylic acid ester with ionic acid groups (including Αί'Αί® and ΕυΌΡΑΟΙΤ®), such as ΕυΌΡΑΟΙΤ ® b 30Ό, ΕυΌΡΑΟΙΤ® РΌ 30Ό, ΕυΌΡΑΟΙΤ® Ρ8 30Ό, ΕυΌΡΑΟΙΤ® b 100-55 and ΕυΌΡΑΟΙΤ® b 30Ό55. In one embodiment of the invention, the pharmaceutical compositions comprise a polymer composition based on proanthocyanidin and an enteric coating, which is a polymer ΕυΌΡΑΟΙΤ® L 30Ό, an anionic copolymer of methacrylic acid and methyl acrylate with an average molecular weight of 250,000 Da. In another preferred embodiment of the invention, the enteric coating coating, which is a polymer, is ΕυΌΡΑΟΙΤ® L 30Ό-55.

- 19 023814

The decomposition of the enteric coating occurs either by hydrolysis by intestinal enzymes, or by emulsification and dispersion by the action of bile salts, depending on the type of coating used. For example, esterases hydrolyze butyl stearate ester to butanol and stearic acid, and since butanol dissolves, stearic acid exfoliates from the drug. Additionally, bile salts are transferred into an emulsion state and dispersed ethyl cellulose, hydroxypropyl methyl cellulose, fats and derivatives of fats. Other types of coatings are removed depending on the time of contact with moisture, for example, coatings obtained from powdered carnauba wax, stearic acid and plant fibers of agar and elm bark are destroyed after plant fibers absorb moisture and swell. The time required for decomposition depends on the thickness of the coating and the ratio of plant fibers to wax.

The enteric coating of the proanthocyanidin polymer can be performed by any method known in the field of enteric coating. For example, but not by way of limitation, enteric polymers can be applied using solutions based on an organic solvent containing from 5 to 10% w / w. polymer when applied by spraying and up to 30% wt./wt. polymer when applied using a drum. Solvents that are commonly used include, but are not limited to, acetone, mixtures of acetone and ethyl acetate, mixtures of methylene chloride and methanol, and ternary mixtures containing these solvents. Some enteric soluble polymers, such as copolymers of methacrylic acid and methacrylic ester, can be applied using water as a dispersant. The volatility of the solvent system must be specially selected so as to prevent sticking due to viscosity and to prevent the formation of high porosity of the coating due to premature spray drying or polymer precipitation as the solvent evaporates.

In addition, plasticizers can be added to the enteric coating to prevent cracking of the coating film. Suitable plasticizers include low molecular weight phthalic acid esters such as diethyl phthalate, acetylated monoglycerides, trityl citrate, polyethylene glycol, tributyl citrate and triacetin. Typically, plasticizers are added at a concentration of 10 wt.% Polymer, which is an enteric coating, and determined by chromatography. Other additives, such as emulsifiers, for example detergents and simethicone, and powders, for example talc, can be added to the coating to increase the strength and smoothness of the coating. Additionally, colorants may be added to the coating to color the pharmaceutical form.

In particular embodiments of the invention, pharmaceutical compositions containing proanthocyanidin-based polymer compositions are provided in the form of enteric coated beads in hard gelatin capsules. Balls of the proanthocyanidin polymer are obtained by mixing a polymer composition based on proanthocyanidin with hydroxypropyl methylcellulose and layering the mixture on nonpareli grains (sugar spheres). In a more preferred embodiment, the directly compressible crofelemer, which does not contain any excipients, additives and media, except for enteric coating, is crushed and divided into balls (for example, such as balls that do not contain non-sugar grains). The balls can be coated with a protective coat of Oraigu C1eag (mixed with water). A preferred enteric coating of beads is ΕυΕΛΟΙΤ ™ Ь 30Ό or ΕυΌΚΛΟΙΤ ™ Ό 30Ό-55, used as an aqueous dispersion containing 20-30% w / w. dry polymer substance, which is provided by emulsifiers lauryl sodium sulfate ΝΡ (8L8) in an amount of 0.7% and polysorbate 80 ΝΡ (Tteeep ™ 20) in an amount of 2.3%, plasticizers such as polyethylene glycol and / or esters are added to this coating citric acid to increase the elasticity of the coating, and talc can be added to prevent the polymer used in the enteric coating from tending to stick together during application and to increase the smoothness of the coating film.

In a specific form, the finished bead composition of the enteric-coated proanthocyanidin-based polymer composition containing nonpareli grains contains 17.3% w / w. nonpareli grains, 64.5% w / w a polymer composition based on proanthocyanidin, 1.5% wt./wt. hydroxypropyl methyl cellulose, 0.5% w / w. Oraigu s1eag, 14.5% w / w ΕυΌΚΛΟΙΤ ™ b 30Ό, 1.45% w / w triethyl citrate and 0.25% w / w. glyceryl monostearate. The specified pharmaceutical form can be obtained by any method known in this field.

A particular form, which is balls of a polymer composition based on proanthocyanidin, does not contain nonpareli grains, contains 78% wt./wt. directly compressible polymer composition based on proanthocyanidin, 0.76% wt./wt. Oraigu s1eag, 19% w / w ΕυΌΚΛΟΙΤ ™ b 30Ό-55, 1.9% w / w triethyl citrate and 0.34% w / w. glyceryl monostearate. The specified pharmaceutical form can be obtained by any method known in this field.

Another form contains 54.58% w / w. balls of a polymer composition based on proanthocyanidin (not containing nonpareli grains and prepared from directly compressible polymer

- 20 023814 compositions based on proanthocyanidin), 1.78% wt./wt. Oraigu s1eag, 39% w / w ΕυΌΚΆΟΙΤ ™ b 30Ό-55, 3.9% w / w triethyl citrate and 0.74% w / w. glyceryl monostearate.

In another embodiment, a pharmaceutical composition comprising a proanthocyanidin-based polymer composition is prepared in the form of granules or powder (microspheres with a diameter of 300-500 p) with an enteric coating, provided in hard gelatin capsules or suspended in an oral solution intended for use by children. Enteric-coated powder or granules may be mixed with food, especially when taken by children. Said preparation may be carried out using methods well known in the art.

In general, granules and powder can be prepared using any method known in the art, such as, but not limited to crystallization, spray drying, or any grinding method, preferably using a high speed granulator mixer. Examples of high-speed granulator mixers include the ΈΙΤΤΕΕΡΟΚΌ ΕΟΌΙΟΕ ™ mixer, the ΕΙΤΤΕΕΡΟΚΌ ΕΟΌΙΟΕ ™ mixer granulator, and the SKL ™ mixer granulator. During mixing of the powder with intense mechanical action, solutions of granulating agents, called binders, are sprayed onto the powder to cause the powder particles to stick together, thereby obtaining larger particles or granules. Granulating agents that are used to prepare the granules include, but are not limited to, cellulose derivatives (including carboxymethyl cellulose, methyl cellulose and ethyl cellulose), gelatin, glucose, polyvinylpyrrolidone (PVP), starch paste, sorbitol, sucrose, dextrose, patcose, acacia gum, sodium alginate, Irish moss extract, panwar gum, ghatti gum, vegetable glue from husks of isapgol seeds, Uedit and arabogalactan from larch wood, polyethylene glycol, ethyl cellulose and waxes. Granulating agents can be added in concentrations ranging from 1 to 30% by weight of particles or granules.

The powder or granules are preferably coated using fluidized bed equipment. Then the granules or powder can be coated with a protective coat of Oraigu C1eag (mixed with water). A preferred enteric coating is ΕυОКЛСИ ™ Ό 30Ό, used as an aqueous dispersion containing 30% w / w. dry polymer substance, which is provided by emulsifiers lauryl sodium sulfate ΝΕ (§6§) in an amount of 0.7% and polysorbate 80 ΝΡ (Feteei ™ 20) in an amount of 2.3%, plasticizers such as polyethylene glycol and esters are added to this coating citric acid, to increase the elasticity of the coating, and talc to prevent the tendency of the polymer used in the enteric coating to stick together during application and to increase the smoothness of the coating film. In one embodiment of the invention, the finished composition in the form of an enteric-coated powder comprises 81.8% w / w. a polymer composition based on proanthocyanidin, 1.5% wt./wt. hydroxypropyl methyl cellulose, 0.5% w / w. Oraigu s1eag, 14.5% w / w ΕυΌΚΑΠΙΤ ™ b 30Ό, 1.45% w / w triethyl citrate and 0.25% w / w. glyceryl monostearate. The finished composition in the form of enteric-coated granules contains 81.8% w / w. a polymer composition based on proanthocyanidin, 10% wt./wt. polyvylpyrrolidone, 1.5% w / w. hydroxypropyl methyl cellulose, 0.5% w / w. Oraigu s1eag, 14.5% w / w ΕυΌΚΑΏΤ ™ b 30Ό, 1.45% w / w triethyl citrate and 0.25% w / w. glyceryl monostearate.

Granules or enteric-coated powder particles may be further weighed in an oral solution, especially for use by children. The suspension can be prepared from aqueous solutions, to which thickeners and protective colloids are added to increase the viscosity of the solution to prevent rapid settling of powder particles or coated granules. As a thickener, any material can be used that increases the degree of hydration of the layer formed around suspended particles through molecular interactions, and which is pharmaceutically compatible with a polymer composition based on proanthocyanidin, such as, but not limited to, gelatin, natural gums (for example, tragacanthane, xanthan, guar, acacia, panwar gum, ghatti gum, etc.) and cellulose derivatives (e.g. sodium carboxymethyl cellulose, hydroxypropyl cellulose and hydroxy propyl methylcellulose, etc.). To improve the action of the thickening agent, a surfactant such as Fetey ™ may optionally be added. A preferred particulate solution comprises a hydroxypropyl methyl cellulose solution in an amount of 2% w / w. in water containing 0.2% Pueep ™.

A polymer composition based on proanthocyanidin can also be prepared in the form of enteric-coated tablets. In one preferred embodiment of the invention, the proanthocyanidin-based polymer composition is granulated using any pharmaceutically acceptable diluent (such as those listed above) by the granule preparation methods described above. The granules are then compressed into tablets using any method well known in the art, for example, but not limited to the wet method

- 21 023814 granulation, dry granulation method or direct compression method. Preferred diluents include, but are not limited to, microcrystalline cellulose (LUGSE ™ PH 200/300) and dextrates (ΕΜΌΕΧ ™). In addition to the tablet form, disintegrants such as those described above and lubricants such as those described above can also be added. A preferred tablet form contains 250 mg of a proanthocyanidin-based polymer composition, 7 mg of AC-ΌΙ-δΟΕ ™ disintegrant (cross-linked sodium carboxymethyl cellulose), 1.75 mg of a magnesium stearate lubricant, and the amount of AU1CE ™ PH 200/300 required to bring the mixture to 350 mg. The tablets are coated using a mixture containing an enteric coating prepared from 250 g of ΈυΌΚΑΟΙΤ ™ L 30 Ό-55, 7.5 g of triethyl citrate, 37.5 g of talc and 205 g of water. The specified form can be obtained by any method known in this field.

In one embodiment of the invention, a directly compressible proanthocyanidin-based polymer composition is prepared in the form of granules by grinding (for example, as described above) and mixed with a lubricant, preferably magnesium stearate. The lubricated granules are then compressed into tablets using any method well known in the art, for example, but not limited to, a direct compression method. Preferably, the weight of each tablet is 125 mg, containing 99.6% w / w. directly compressible polymer composition based on proanthocyanidin and 0.40% wt./wt. magnesium stearate. Then the tablets are preferably coated with a mixture containing an enteric coating, said mixture containing 30% suspension (6.66 g of 22.22 g) ΕυΌΚΑΟΙΤ ™ L 30Ό-55, 0.67 g of triethyl citrate, 1.67 g of talc and 20.44 g of purified water per 100 g of tablet. Tablets may be prepared by any method known in the art.

In a more preferred embodiment of the invention, a directly compressible polymer composition based on proanthocyanidin is prepared in the form of inner layers of tablets weighing 125, 250 or 500 mg, containing 99.6% w / w. directly compressible polymer composition based on proanthocyanidin and 0.40% wt./wt. magnesium stearate. The tablets are then preferably coated with a mixture containing an enteric coating. The finished tablet composition contains 86.6% wt./wt. directly compressible polymer composition based on proanthocyanidin, 0.4% magnesium stearate, 6.5% ΕυΌΚΑΟΙΤ ™ 30Ь-55, 0.9% triethyl citrate, 2.87% talc and 2.74% white dispersion. Tablets may be prepared by any method known in the art, for example, by the method described below, but not limited to.

Compositions obtained in the form of microparticles (which include particles of the order of several micrometers, such as microspheres and microcapsules), particles (which include particles of the order of several millimeters), crystals of drugs, pellets, pills and beads, can be coated using the method spray coating in a fluidized bed. In this method, fluidized bed equipment is used, such as supplied by OATT ™, AEKOMAT1C ™, \ υΚδ, ™ or other companies, in which the inner layers of the composition are carried upward in a closed cylindrical vessel using a stream of air coming from below, and an enteric layer is obtained by spray drying this layer on the inner layers during fluidization. To coat tablets or capsules with a coating, Asse1a-Co! A (MAZHU ™) coating equipment can be used. According to the method, tablets or capsules are placed in a rotating cylindrical coating drum with a perforated casing, and spraying devices are placed inside the drum and dry air is passed through the rotating tablets or capsules. Any other type of coating drum may also be used, such as a COMRE-LAB ™ drum. Ηίsoa! Ο ^ ΑΤΤ ™ with submersible sword processing, EpasoYug EC1AM ™, 8ΤΕΙΝΒΕΚΟ ™ equipment, ΡΕΕΕΕΟΚΙΝΙ ™ equipment or ΑΑΙ, ΤΙ ΙΚΚ 'equipment.

In another embodiment, the proanthocyanidin-based polymer composition is provided in the form of a suppository for rectal administration. Suppositories can be prepared using any basic substance that is pharmaceutically acceptable for the manufacture of suppositories and compatible with the polymer composition based on proanthocyanidin. Since rectal administration does not expose the proanthocyanidin-based polymer composition to the stomach environment, pharmaceutical forms for rectal administration need not be prepared to protect the composition from the stomach environment. The bases that can be used to prepare suppositories containing a polymer composition based on proanthocyanidin include, but are not limited to, cocoa butter, gelatin with glycerin, hydrogenated vegetable oils, mixtures of polyethylene glycols or fatty acids of polyethylene glycols, or a combination of glycols and surface active substances, or non-ionic surfactants (such as polyoxyethylene sorbitan fatty acid esters (Τ ^ ββηδ), polyoxyethylene stearates and mixtures of sorbitan esters fatty acids (and 8rap Ag1ase1)). However, due to the hydrophilicity of the proanthocyanidin-based polymer composition, hydrophilic bases for suppositories are proposed. Preferred

- 22 023814 a form of a polymer composition based on proanthocyanidin, which is a suppository, is prepared from 91 g of glycerol, 9 grams of sodium stearate, 5 g of purified water, this form may contain from 5 to 50% wt./wt. polymer composition based on proanthocyanidin. Alternatively, the suppository may contain 10 g of a polymer composition based on proanthocyanidin, 20 g of gelatin and 70 g of glycerin. Suppositories made from a proanthocyanidin-based polymer composition may be formed by any method known in the art, including but not limited to compression molding, melting, or preferably melting in the mold.

In another embodiment, a proanthocyanidin-based polymer composition is prepared using a compound or compounds that neutralize gastric acid. Alternatively, a pharmaceutical composition comprising a proanthocyanidin-based polymer composition is administered simultaneously with or after administration of a pharmaceutical composition that neutralizes gastric acid. Compounds, such as antacids that are used to neutralize stomach acid, include, but are not limited to, aluminum carbonate, aluminum hydroxide, bismuth subnitrate, bismuth subsalicylate, calcium carbonate, sodium dihydroxyaluminium carbonate, magaldrate, magnesium carbonate, magnesium hydroxide, magnesium oxide and mixtures thereof.

In another embodiment, the proanthocyanidin polymer composition is administered with a substance that inactivates or inhibits the action of stomach enzymes such as pepsin. Alternatively, a pharmaceutical composition comprising a proanthocyanidin-based polymer composition is administered simultaneously with or after administration of the pharmaceutical composition to inactivate or inhibit the action of gastric enzymes. For example, but not by way of limitation, protease inhibitors such as aprotin can be used to inactivate gastric enzymes.

In another embodiment, a proanthocyanidin-based polymer composition is prepared using a compound or compounds that inhibit the secretion of gastric acid. Alternatively, a pharmaceutical composition comprising a proanthocyanidin-based polymer composition is administered simultaneously with or after administration of the pharmaceutical composition to inhibit gastric acid secretion. Compounds that are used to inhibit the secretion of gastric acid include, but are not limited to, omeprazole, esomeprazole, lansoprazole, rabeprazole, pantaprazole, ranitidine, nizatidine, famotidine, cimetidine and misoprostol.

The following series of examples are provided for illustration and do not limit the scope of the invention. Experimental Section

Comparative example. Isolation of a partially purified proanthocyanidin-based polymer composition.

The directly compressible polymer composition based on proanthocyanidin was isolated from the milk juice of the CGyOp 1ESYeP plant as follows: 460 L of the milk juice of CGYOUP 1ESYeP was mixed with 940 L of purified water for ten minutes, and then insisted overnight (12 h) at 4 ° C. The red supernatant was pumped into the holding tank, and the sediment was removed. Then, the supernatant was extracted with 200 L of n-butanol by stirring for ten minutes, and then phase separation was achieved. The n-butanol phase was removed, and the aqueous phase was extracted two more times, each time with 200 L of n-butanol. After extraction, the aqueous phase was concentrated by ultrafiltration using a membrane with a cut-off level of 1 kDa (low protein binding of the cellulose membrane), and then the retentate was dried in a centrifugal dryer at a temperature of about 37 ° C (± 2 ° C).

For purification by column chromatography, 6 kg of the dried extract was dissolved in 75 L of purified water and stirred for 90 minutes. The dissolved material was subjected to chromatography in a system of two chromatographic columns, consisting of a 35-liter column CM-§perbato8e (weakly acidic cation exchange resin) and a 70-liter LN-20 column (resin for size exclusion chromatography) connected in series. The material was loaded onto a SM-Apolato8e column, washed with 140 L of purified water, and then washed into a LN-20 column with 375 L of 30% acetone. At this stage, the two columns were separated and the proanthocyanidin-based polymer composition was washed from the LH20 column with 250 L of 45% acetone. Fractions were collected in 10-liter containers and monitored using a UV detector at 460 nm. Fractions containing absorption material at 460 nm were pooled and concentrated by ultrafiltration using a membrane with a cut-off level of 1 kDa (low protein binding of cellulose membrane). The retentate was dried using a rotary evaporator in a water bath at a temperature of about 37 ° C (± 2 ° C) to obtain a partially purified polymer composition based on proanthocyanidin.

A partially purified proanthocyanidin-based polymer composition was tested for direct compression. A portion of a 250 mg proanthocyanidin-based polymer composition in the absence of any binders or excipients was placed in a tablet machine and then compressed into tablets of various thicknesses (for example, the greater the pressure on the composition to obtain a tablet, the smaller the resulting tablet) . Then, using the hardness tester, the strength of the tablets was determined.

- 23,023,814

The abrasion of tablets with a strength of 8-15 kp was determined as described in IPR 23 (1216). Abrasion was less than 0.5% weight loss.

Example 1. Obtaining a polymer composition based on proanthocyanidin.

The partially purified proanthocyanidin-based polymer composition obtained according to the comparative example was weighed and ground in a glass beaker using a glass rod. The crushed polymer composition was dissolved and titrated with 0.5 N. ΝαΟΗ solution to pH 8-8.1, and then treated with dichloromethane six times. The aqueous layer was exposed to a bath at a temperature of 60 ° C without applying vacuum to remove any amount of dichloromethane remaining. The residue was titrated to pH 4 using 1N. HC1 solution. A sufficient amount of acetone was added to obtain a 10% acetone solution. The bH-20 resin was balanced with distilled water at pH 4, placed in a Buchner funnel, and excess liquid was removed. The aqueous acetone residue containing the proanthocyanidin polymer was mixed with an equal amount of semi-dried LH-20 resin and allowed to settle for 30 minutes. LN-20 in an amount equal to the initial aliquot of LN-20, previously adjusted in the same way, was filtered on a Buchner funnel through filter paper ΧνΐιαΙιηαη 42 to remove excess liquid. The combination of LH-20 and the proanthocyanidin polymer was placed on top of the LH-20 layer in a Buchner funnel and allowed to infuse for 30 minutes. After settling 140 ml of 10% wt./wt. A solution of acetone in deionized water at pH 4 was added to the top of the Buchner funnel. A slight vacuum was created to begin collecting the eluate. This operation was repeated 5 more times to obtain a total of 6 fractions of 140 ml each. A total of 840 ml of solvent with 10% w / w was used. by washing. Additional leaching was carried out using 25% wt./wt. acetone solution (10x140 ml), 32% w / w. acetone solution (6x140 ml), 39% w / w. acetone solution (6x140 ml), 45% w / w. acetone solution (6x140 ml) and 54% w / w. acetone solution (6x140 ml). The fractions containing the proanthocyanidin polymer were combined and dried in a vacuum oven at a temperature of 60 ° C to obtain a polymer composition based on proanthocyanidin with a yield of 16.59%. A comparison of the purity of the composition of Example 1 with that of the comparative example is shown in the following table.

Comparative example and example 1

Cleanliness defined by HPLC Taspine content Comparative Example 63% 0.4% Example 1 92% 0.1%

Alternatively, the columns could be split and did not work sequentially. An example of how split columns work is as follows. For example, columns could be run as two or more separate objects. For example, 6 kg of the solute was loaded into the first column (for example, CM-Speryago 8e) and solution A was collected. Since the column was washed with water, solution B was collected. Then, since the column was washed with approximately 30% acetone solution, solution C was collected.

Then, these solutions (solutions A, B and C) were sequentially (for example, solution A, then solution B, and then solution C) loaded into a second column (for example, LN-20). Then, crofelemer was washed out of the column (e.g., LH-20) with approximately 45% acetone solution.

Example 2

Step 1. The same volume of methyl ethyl ketone was added to the crude solution of plant milky juice (500 ml), the mixture was carefully mixed and kept without moving for 2 hours. The organic layer was separated and the process was repeated two times. The aqueous layer was concentrated to obtain 60 g of the composition in step A.

The size servings Layer Exit (%) Crofelemer (purity%) Taspin (parts per million) 50 ml Water thirteen 65.9 45178 50 ml Water thirteen 62.0 42026 50 ml Water thirteen 63.3 42704 0.5 l Water 12 64.3 55141 100 l Water thirteen 77.2 35039

Stage 2. 10 g of the composition obtained in stage A, was dissolved in 120 ml of water and stirred for 1 h at a temperature of 30-35 ° C, and then kept at room temperature for another 1 h. The mass was filtered through filter paper in a vacuum and the filtrate was exposed in an 8eryago 8e column (11 cm, inner diameter 3 cm), connected in series with a Zeriyeh column (22 cm, inner diameter 3 cm). Washing was carried out with 250 ml of water, then with a 30% solution of acetone in water (600 ml). After washing with a 30% acetone solution, the Zeryagoke column was separated and the column was washed with an 8perFe \ 45% solution of acetone in water. The first 110 ml of the fraction was discarded, and then 100 ml of the dark-colored eluate was collected and concentrated to provide 1.3-1.5 g of crofelemer.

The size servings Exit (%) Crofelemer (purity%) Taspin (parts per million) Mp Ρϋ South 14 91.0 NGOs 2586 1.41 South 12 96.0 NGOs 2700 1.41

* NGOs - below the detection limit; * MP - molecular weight, * ΡΌ - polydispersity.

Example 3

Stage 1. Two volumes of methyl ethyl ketone were added to the crude plant milk juice solution (500 ml) containing a precipitate, gently mixed and kept without moving for 2 hours. The organic layer was separated and the process was repeated three more times. The combined organic layers were concentrated to give 50 g of the composition in step A.

The size servings (liters) Layer Exit (%) Crofelemer (purity%) Taspin (parts per million) 0.2 IEC 10 54.1 9608 0.2 IEC nine 47.0 12465 0.2 IEC 8 69.8 16571 0.5 IEC 10 53.5 134281 2.0 IEC 8 61.7 16612

Stage 2. 50 g of the composition obtained in stage A, was dissolved in 10 vol. distilled water and stirred for 1 h at a temperature of 30-35 ° C. The solution was kept at room temperature for an additional 1 h. The solution was exposed in a Spergo8e column (6 vol.). Washing was carried out 30 vol. water, then 30 vol. a solution of acetone in water. Washed by water and 8 vol. organic mixture (30% acetone) was discarded, followed by washing out the dark eluate, fractions with a high product content (15 vol). A fraction with a high product content was selected and concentrated to provide a yield of crofelemer of 7.5-8.0 g.

The size servings (g) Exit (%) Crofelemer (purity%) Taspin (ppm) Mp Ρϋ fifty sixteen 94.5 NGOs 2373 1.31 fifty sixteen 96.0 NGOs 2384 1.39

Example 4

Stage 1. To the crude solution of plant milky juice (500 ml) containing a precipitate was added 2 vol. methanol and four vol. ethyl acetate, stirred for 4 hours and kept motionless for 2 hours. The organic layer was separated and filtered through filter paper. The filtrate was concentrated to obtain 90 g of the composition in step A.

The size servings (liters) Exit (%) Crofelemer (purity%) Taspin (parts on million) 0.5 eighteen 64.6 27837 0.5 18,4 65.11 28181 0.5 17 65.09 27812

Stage 2. 50 g of the composition obtained in stage A, was dissolved in 12 vol. distilled water and stirred for 1 h at a temperature of 30-35 ° C. The solution was kept at room temperature for an additional 1 h. The solution was exposed in a Spergo8e column (6 vol.). Washing was carried out 30 vol. water, then 25 vol. a solution of acetone in water. Washed by water and 8 vol. the organic mixture (30% acetone) was discarded, followed by washing out the dark eluate, fractions with a high product content (15 vol.). A fraction with a high content of the product was selected and concentrated to provide a yield of crofelemer of 12.0-13.0 g.

The size servings (D) Exit (%) Crofelemer (purity%) Taspin (parts on million) Ml Ρϋ fifty 26 98.25 NGOs 2032 1,03 fifty 25,4 98.64 NGOs 2417 1.01 fifty 24.6 100.9 NGOs 2401 1.01

- 25,023,814

Example 5

A method for purifying crofelemer by treating with charcoal in methanol / ethyl acetate by a precipitation method.

The untreated proanthocyanidin compound (100 g) was dissolved in methanol (1000 ml) and filtered through a sintered glass funnel. Insoluble solids were re-extracted with methanol (200 ml), followed by combining the two filters, and the combined filtrates were treated with 30 g of charcoal with stirring for 60 minutes. The compound was filtered twice through a layer of bentonite, and the residue was removed. The filtrate was treated with 6.2 ml of ethyl acetate and stirred for 60 minutes, after which it was passed through a 2 mm thick layer of paper \ νΐι; · ιΙιη; πι. Received 52 g of a brown solid with a purity of 99.81% and a taspine impurity content below the detection limit.

The use of bentonite to remove protein in an aqueous solution.

3.3520 g of a partially purified proanthocyanidin-based polymer composition was weighed and placed in a plastic tube with a 50 ml cap; 30 ml of deionized water (the pH of which was previously adjusted to pH 8.8) was added to dissolve the entire partially purified polymer composition with shaking. The pH of the resulting solution was 6.12 and was adjusted to 5.5 with 1N. HC1 solution. Then the solution was evenly distributed over two tubes (a certain amount of foam remaining in the original tube was removed).

One of the tubes was shaken for 1 h, and then kept at room temperature for 22 h. The resulting solution was filtered through a funnel with a fine mesh filter. A 100 μl sample of the filtrate was recovered, dried to give 9.7 mg of solid, and tested by amino acid analysis for use as an untreated bentonite sample subjected to AKA. The pH of the filtrate was adjusted to pH 8.0 with 0.5N. NaOH solution. 50 μl of the sample was removed, dried and tested by HPLC and the molecular weight distribution of the sample not treated with bentonite was determined.

600 mg of bentonite was added to the second tube. The tube was closed and shaken for 1 h, and then kept at room temperature for 22 h. The resulting solution was filtered through a funnel with a fine mesh filter. A 100 μl sample of the filtrate was recovered, dried to obtain 10.3 mg of solid, and tested by amino acid analysis for use as a bentonite-treated AKA sample. The pH of the filtrate was adjusted to pH 8.0 with 0.5N. NaOH solution. 50 μl of the sample was removed, dried and tested by HPLC and the molecular weight distribution of the sample treated with bentonite was determined.

The use of ethyl acetate to treat a partially purified polymer composition.

1.6704 g of the partially purified polymer composition was weighed and slowly added to a scintillation vial with a stirrer for a magnetic stirrer and 15 ml of deionized water (the pH of which was previously adjusted to pH 8.62). The vial was closed and the contents were mixed until the entire amount of the partially purified polymer composition was dissolved. The pH of the resulting solution was 6.49, and then brought to 8 by 0.5N. NaOH solution. 200 μl of the sample was recovered, dried and tested by HPLC and the molecular weight distribution of the sample was determined before extraction by EJAc.

The solution of the partially purified polymer composition, the pH of which was 8, was extracted six times with 20 ml of EAAc. During extraction, similar to extraction with dichloromethane, the formation of an interfacial layer was noted. 200 μl of the sample was recovered from the obtained aqueous phase, dried and tested by HPLC, and the molecular weight distribution of the sample was determined after extraction by EJAc.

The use of bentonite and MeOH / ENAc for the processing of materials of a partially purified polymer composition.

Serving Size 2.5 g.

2.5 g of the partially purified polymer composition was placed in a 50 ml polypropylene tube with a lid and 25 g (~ 32 ml) of MeOH was added. The tube was closed and placed in a shaker for shaking for 10 min until all solids were dissolved. A sample of this solution in an amount of 130 μl was dried in an oven under vacuum at room temperature (CT) and subjected to testing.

750 mg of bentonite was added to a solution of the partially purified polymer composition in methanol, and then the tube was closed and shaken for 1 h. The solution was filtered through filter paper using a Buchner funnel on an Erlenmeyer flask. Bentonite together with the material adsorbed on it was removed. 130 μl of sample was recovered from the filtrate, dried under vacuum in an oven at room temperature, and then analyzed.

The above filtrate (32 ml) was placed in a 250 ml polypropylene bottle, after which a five-fold volume of EyAc (160 ml) was added. An immediate brown precipitate was detected. The bottle was closed and shaken for 10 minutes.

The mixture was filtered through filter paper using a Buchner funnel. The solid was dried under vacuum at room temperature to obtain 930.2 mg of the product; a sample of the product was analyzed. A certain amount of particles passed through the filter paper and penetrated into the filtrate. The indicated filtrate was centrifuged and the precipitated solid was filtered off and washed with ЕУАс, dried under vacuum at room temperature to obtain another 194.8 mg of solid. A sample was taken from the indicated solid. The pure filtrate and the EJAc wash solution were combined and dried by rotary evaporation at 40 ° C. to obtain 538.7 mg of a solid. A sample was also taken from the indicated solid.

Serving Size 12.5 g.

The method described above was carried out using an increased amount of partially purified polymer composition, amounting to 12.5 g. After addition of ЕУАс with precipitation of the desired products, the mixture was filtered through filter paper. The solid over the filter paper was dried under vacuum at room temperature to obtain 4.4896 g of solid. A sample was taken from the indicated solid. Similar to the smaller scale method, the filtrate still contained some solid, and the filtrate was left in the refrigerator overnight. Then, the resulting solution containing the precipitate was slowly poured into a Buchner funnel through filter paper, this time the solid did not pass through the filter paper. The remaining solid was dried in vacuo at room temperature to give 1.4221 g of solid. A sample of the substance was taken.

Studying protein removal with a 2.5 g serving size

Previously, samples for amino acid analysis before and after treatment with bentonite were not taken, the experiments were repeated and the indicated volume of solutions, which are samples for amino acid analysis, were removed before and after treatment with bentonite and dried. After treatment with bentonite, the solution was filtered before sampling. To exclude the effect of protein removal only by filtration, before treatment with bentonite, the solution was also filtered before sampling. Thus, the difference in protein levels in these two samples should be associated with treatment with bentonite.

Obtaining pharmaceutical forms.

The invention describes illustrative methods for the production and packaging of pharmaceutical forms of a polymer composition based on proanthocyanidin obtained from C.1ecyep according to the methods proposed in the invention.

Enteric-coated balls encapsulated.

The following are descriptions of the methods used to produce a proanthocyanidin-based polymer composition in the form of an enteric-coated enteric-coated bead based on sugar spheres. Each hard gelatin capsule contained 250 mg beads of an enteric-coated proanthocyanidin polymer composition. The capsules were packaged in NIR bottles containing sixteen (16) capsules weighing 250 mg each. The enteric coated proanthocyanidin-based polymer composition beads contained 17.3% (w / w) nonpareli grains (40/60 mesh sugar spheres, Rayaig, series No. 60084060), 64.5% of the polymer composition per based on proanthocyanidin from Cl.1ecy1ep, 1.5% hydroxypropylmethyl cellulose (Me1ose1 E5 Prgetst, Iote Syetuya1 Co, series no. MM9410162E), 0.5% OraFu Cleag (Co1ogsop, series no. 883563), 14.5% EiOpOKH ™ These., Batch No. 1250514132), 1.45% triethyl citrate (Mogyekh, series No. N5X291), glyceryl monostearate (1ш No. Yog-900, Koit Tes, series No. 502-229) and purified in ode (И8Р).

A coating solution containing a proanthocyanidin-based polymer composition was prepared by adding hydroxypropyl methylcellulose and a proanthocyanidin-based polymer composition to purified water (I8P) and mixing until completely dissolved. Nonpareli grains were loaded into the chamber of a fluidized-bed dryer (Precision Coating Unit No. G). Then, the polymer solution was applied in layers to non-paired grains by spraying the solution onto fluidized non-paired grains at a target bed temperature of 3035 ° C. After the deposition of the proanthocyanidin polymer layers was completed, a protective coating was applied using OraFgu C1eag (prepared by mixing OraFu C1eag with purified water, I8P) at a target layer temperature of 30-35 ° C. After the protective coating was applied, the pellets were removed and sieved through a membrane with parameters 1000 1 and 425 μ, and layered spheres, the parameters of which are more than 425 μ and less than 1000 μ, were poured back into the fluidized bed dryer. Meanwhile, an enteric-coating solution was prepared by mixing triethyl citrate and glyceryl monostearate with water, which was heated to 65 ° C, and then mixing the solution with EuCAO1T ™ 30Ό-55. The resulting enteric coating solution was then sprayed onto layered spheres in a fluidized bed dryer at a bed temperature of 30-35 ° C. until the entire enteric coating solution was applied to the pellets. Enteric coated beads were manually poured into hard gelatin capsules of size 0 to provide a dose of 250 mg, and then

- 27 023814 were packaged in suitable bottles ΗΌΡΕ with a lid sealed from the inside using induction heating.

Serving composition.

Product: enteric-coated proanthocyanidin polymer beads.

Serving Size: 578.0 g.

Raw materials: quantity per serving.

Sugar spheres, nonparelle, ΝΕ (40/60): 100.0 g.

Proanthocyanidin-based polymer composition: 372.8 g.

Hydroxypropyl methylcellulose E5, υδΡ (K29 / 32): 8.7 g.

Orabgu C1eag (U5-1-19025A): 2.9 g.

ΕϋϋΚΑΟΠ ™ L 30Ό-55 (30% solids): 279.4 g.

Triethyl citrate, ΝΕ: 8.4 g.

Glycerin monostearate: 1.4 g.

Water, υδΡ (removed during processing): 1284.8 g.

Enteric-coated balls encapsulated.

The following describes methods for producing enteric coated beads enclosed in capsules that do not contain nonpare sugar spheres. One form contains 83.3% w / w. a polymer composition based on proanthocyanidin, 0.5% wt./wt. Orabgu s1eag, 14.5% w / w EINKASHT ™ L 30Ό-55, 1.9% w / w. triethyl citrate and 0.34% glyceryl monostearate.

The balls were coated with the first protective layer using a 5% solution of Orabgu s1eag in a 16 liter Aetotais MP-1 dryer with a fluidized bed and a 50 mm Wurster column. The parameters for applying the protective coating were the inlet temperature from 50 to 60 ° C, the outlet temperature from 25 to 40 ° C, the volume of transported air from 30 to 40 m ³ / h, the spray rate from 6 to 12 g / min and the air pressure 2.5 bar. After the protective coating was applied, the balls were removed and sieved to select balls with parameters greater than 425 μ and less than 10001.

Then, suitable sized beads were poured back into the fluidized bed dryer for enteric coating. For every 1000 g of proanthocyanidin-based polymer composition balls, a suspension of enteric coating was prepared from 811.97 g of EIB ™ 30Ό-55, 24.36 g of triethyl citrate, 4.36 g of glyceryl monostearate and 248.55 g of purified water. The specified suspension was prepared by continuously carefully mixing the suspension EEONAOP ™ L 30Ό-55, as well as obtaining suspension and homogenization in a separate container of triethyl citrate and talc in purified water. Then a mixture of triethyl citrate and talc was added to a suspension of EIϋΚΑΟΓΤ ™ L 30Ό-55 and the resulting coating dispersion was mixed during spraying to avoid settling. Then the balls were coated in a fluidized bed dryer with the following parameters: inlet temperature ranged from 42 to 47 ° C, outlet temperature ranged from 28 to 34 ° C, air flow rate was 30-40 m ³ / h, spray rate was 6 -12 g / min and the air pressure was 2.5 bar. Then, the resulting enteric-coated beads were poured into No. 0 capsules with hard gelatine shells.

Enteric-coated granules and powder.

The following is a method for preparing a polymer composition based on proanthocyanidin in the form of enteric-coated granules or powder, enclosed in hard gelatine capsules or suspended in an oral solution. The particles of the powder of the polymer composition based on proanthocyanidin were prepared by mixing with intensive mechanical action the powder of the polymer composition based on proanthocyanidin and hydroxypropyl methylcellulose in a high-speed granulator mixer. The granules of the proanthocyanidin-based polymer composition were prepared by spraying polyvinylpyrrolidone onto a powder in a high-speed granulator mixer, so that the powder particles coalesced into larger granules. Then, the granules or powder was coated with a protective layer of Orabgu C1eag (mixed with water) using fluidized bed equipment, and then coated with an enteric-soluble EEPNAOP ™ 30Ό coating applied in the form of an aqueous dispersion containing 30% w / w. dry methacrylate polymer, which was provided by emulsifiers of sodium lauryl sulfate ΝΡ (δΗδ) in an amount of 0.7% and polysorbate 80 ΝΡ (TGEP ™ 20) in an amount of 2.3%, plasticizers, triethyl citrate and glyceryl monostearate were added to this coating to increase the elasticity of the coating . The finished enteric-soluble powder composition contained 81.8% w / w. a polymer composition based on proanthocyanidin, 1.5% wt./wt. hydroxypropyl methyl cellulose, 0.5% w / w. Orabgu s1eag, 14.5% w / w EEEROI ™ L 30Ό, 1.45% w / w. triethyl citrate and 0.25% w / w. glyceryl monostearate. The finished composition of the enteric-coated granules contained 81.8% w / w. a polymer composition based on proanthocyanidin, 10% polyvylpyrrolidone, 1.5% w / w. hydroxypropyl methyl cellulose, 0.5% w / w. Orabgu s1eag, 14.5% w / w EEEROI ™ L 30Ό, 1.45% w / w. triethyl citrate and 0.25% w / w. glyceryl monostearate.

Granules or particles of a proanthocyanidin-based enteric-coated polymer composition may be poured into hard gelatin capsules in an amount that provides a suitable dose of the drug. Granules or powder particles of an enteric-coated proanthocyanidin polymer composition may also be weighed in an oral solution, especially for use by children. A suspension solution was prepared by soaking 2 g of hydroxypropyl methylcellulose in 97.8 ml of distilled water and 0.2 g of Tteep ™ 80, stirring the preparation to a uniform state by means of ultrasound, heating the solution to 40 ° C and stirring for three hours, and then adding granules or particles of a polymeric composition based on proanthocyanidin with enteric coating to a homogeneous solution.

The following describes a method for preparing a polymer composition based on proanthocyanidin using a diluent in the form of enteric-coated tablets. For each tablet weighing 350, 250 mg of the proanthocyanidin-based polymer composition was granulated using 7 mg of cross-linked sodium carboxymethyl cellulose (AC-E1-8OB ™) and a mass of microcrystalline cellulose (A ^ CEB ™ PH 200/300) sufficient to bring the total weight up to 350 mg. These components were mixed for a period of time ranging from 20 to 30 minutes in a V-shaped mixer. After stirring for 20 to 30 minutes, 1.75 mg of magnesium stearate was added and the mixture was stirred for an additional period of 4 to 5 minutes. The obtained granules were compressed on a rotary tablet press using 5/16 inch standard concave punches. The tablets were coated with an enteric coating mixture prepared from 250 g of ΕυΌΚΑΟΙΤ ™ L 30 Ό-55, 7.5 g of triethyl citrate, 37.5 g of talc and 205 g of water. Then, the tablets were placed in a perforated coating drum (for example, ACCEPACOTA ™ system) and rotated at a speed of 15 rpm./min at a temperature of 40 ° C. The enteric coating was sprayed with the following parameters: the inlet air temperature was 44-48 ° C, the exhaust air temperature was 29-32 ° C, the product temperature was 26-30 ° C, the diameter of the spray nozzle was 1 mm, and the rotation speed of the drum was between 30 to 32 rpm, the air flow rate was 30-32 cubic meters. ft / min and a spray pressure of 20 psi. inch. Finally, the tablets were held for 30 minutes while rotating the drum at a speed of 15 rpm and an inlet air temperature of 60 ° C, and then after stopping the supply of heat, the tablets were rotated at a speed of 15 rpm until the tablets cooled to room temperature.

Enteric coated tablets obtained by direct compression.

A method for preparing a polymer composition based on proanthocyanidin in the form of enteric-coated tablets without the use of a diluent was performed as described below. A polymer composition for direct compression based on proanthocyanidin was obtained. 125 mg tablets were prepared by mixing 99.6% w / w. a polymer composition for direct compression based on proanthocyanidin with 0.40% wt./wt. magnesium stearate for two minutes, followed by direct compression of the material into tablets weighing 125 mg on a rotary press using standard round punches with a concave surface with a diameter of 1/4 inch to achieve a strength of 4-10 kp.

Testing of tablet cores showed that the average strength (n = 10) was 4-10 kp, the abrasion (n = 20) was less than 0.7%, the average tablet weight (n = 10) was 125 mg ± 7 mg, and the average thickness ( n = 10) ranged from 3.9 to 4.1 mm, and disintegration (n = 6) was no more than 20 minutes.

The dispersion for coating was prepared by mixing, while to obtain 100 g of tablets, 22.22 g of 30% wt./wt. Suspensions ΕυΟΚΛΌ ™ L 30Ό-55 were carefully mixed with a mixture consisting of 0.67 g of triethyl citrate, 1.67 g of talc and 20.44 g of purified water, stirring until homogeneous. The coating dispersion was continuously mixed to avoid settling.

The tablets (in portions of 100,000) were coated with a coating dispersion in a Sotri-BaL drum 24 inches / 30 L. The tablets were mixed in a drum at a speed of 3-5 rpm and preheated to a temperature of 35 to 40 ° C. Then, a dispersion of enteric coating was applied to the tablets until a tablet weight gain of 6 to 8% was achieved, with the following parameters: inlet temperature was 45-65 ° C, exhaust air temperature was 27-34 ° C, product temperature was 28-32 ° C, the rotation speed of the drum was from 8 to 14 rpm, the air flow rate was from 180 to 240 cubic meters. ft / min, a spray pressure of 10-20 psi, an initial spray rate of 3 to 4 g / min / kg, and a final spray rate of 4 to 8 g / min / kg. Then the tablet was kept in the drum for 30 minutes at an inlet temperature of 45 to 50 ° C and a drum rotation speed of 3 to 5 rpm. Finally, the tablets were allowed to cool to room temperature in a drum with a rotation speed of 3 to 5 rpm. Then four tablets weighing 125 mg were poured into an opaque gelatin capsule, stained with 8 \\ υάίκ1ι, size 0.

The enteric-coated proanthocyanidin-based polymer composition tablets were tested for uniformity of composition, release and stability, microbiological tests were performed, and some analytical process tests were performed. In stability studies, the proanthocyanidin-based polymer composition remained stable after six months of storage at room temperature, as well as with increasing temperature and humidity. Finally, the inner layer of the tablets was tested and found that the average strength (n = 10) was 4-10 kp, the abrasion (n = 20) was less than 0.7%, the average weight of the tablet (n = 10) was 125 mg ± 7 mg , the average thickness (n = 10) was from 3.9 to 4.1 mm, and the disintegration (n = 6) was no more than 20 minutes.

Enteric coated tablets obtained by direct compression.

Forms of the proanthocyanidin-based polymer composition without diluent described in the invention, which are enteric coated tablets, were made as described below. Tablet cores were prepared by grinding a proanthocyanidin-based polymer composition in an amount of 250 mg per tablet (about 16 kg in total) in Oiaigot SotP with 024K sieve (30 mesh), and then mixing the milled composition in a Rayegkop Ke11eu ν-shaped mixer with a volume of 2 cubic feet. Then, magnesium stearate was added to the composition in the mixer (Zresyit Onu Shu RgoPps1k, Shs., No. Vgipkuyuk, Ν.Τ) in an amount of 1 mg per tablet and stirred for 2 minutes. Then, on a rotary tablet press, the mixture was pressed into tablets weighing 251 mg (containing 250 mg of a polymer composition based on proanthocyanidin) with a strength of 8-15 kp and an abrasion rate of less than 0.5%.

The dispersion of the coating was prepared by first mixing in a first vessel 25 g (7.5 g of solid particles) of a dispersion of EEERACH ™ 30 Ό-55 (NiK Ateis, Sch., ZotegkeE N.1.) (Weight indicated on 115 g of coated tablets ) A dye dispersion was prepared by sequentially adding to the second container with constant stirring 39.59 g of purified water, 3.30 g of talc (A1рйай1 ™ 500) (Shyakeg, S1agk & Oap1e1k, Shs., ZoySh R1atPe1P, Ν.Ρ), 6, 06 g (3.15 g of solid particles) of a white dispersion (coloring material) (^ agpeg-ShikShkop, Shs., Δ. Shoshk, Mo.), and then 1.05 g of triethyl citrate (Mogyekh, Shs., Szhepkogo, ΝΌ. ) The mixture was then homogenized for 15 minutes or until a uniform consistency was achieved. With slow stirring, the dye dispersion was added to the EuERACH ™ 30 30-55 dispersion, and then mixed for 30 minutes before spraying. Also, stirring was continued during the spray treatment to avoid settling.

The tablets were coated with portions of 50,000 tablets in a Sotri-BaL 24 inch / 30 L drum with the following parameters: atomization air pressure was 10-20 psi, air entering the drum was 35-60 ° C, distance from tip the nozzle to the tablet layer was 5 to 6 inches and the number of septa / nozzles was 4/2. After adding tablets, the drum was rotated at a speed of 3 to 5 rpm and was heated to a temperature of 40 ° C. Then, the coating was sprayed on the tablets until a tablet weight gain of 11 to 13% was achieved, with the following parameters: the target temperature of the exhaust air (achieved within ten minutes of spraying) was 27-33 ° C; the rotation speed of the drum was from 8 to 12 rpm, the air flow rate was from 180 to 240 cubic feet / min and the spraying intensity was 2-5 g / min / kg. After achieving the desired mass gain, the heating was stopped and the drum was rotated at a speed of 3-5 rpm until the tablets were cooled to a temperature below 30 ° C. The tablets were enclosed in opaque gelatin capsules ΌΒ stained with Zuehky Ogapde, size AA (Sarkide1, SgeepuooP, Z.S.). Tablets weighing 500 mg were also prepared as described above, except that the coating was prepared per serving of 25,000 tablets with a tablet weight gain of 8 to 10%.

Dry powder for oral suspension.

Polymeric composition based on proanthocyanidin (1.6 g), sodium benzoate (0.06 g), microcrystalline cellulose and sodium carboxymethyl cellulose (A ^ CE Cb 611, 0.04 g), colloidal silicon dioxide (0.01 g), flavoring Tiy1 RGiSh (0.03 g) and pharmaceutical grade sugar (9.25 g) were individually sieved through a No. 40 sieve, mixed in geometric proportion, and then mixed in a mixer. The dry mixture (with a total mass of about 11 g) was a loose powder from light brown to brown in color and had a bulk density of 0.78 g / ml.

The dry mixture, when reconstituted with 23 ml of purified water, formed a brown suspension (approximately 30 ml), the pH of which was 5.2. Every 5 ml of reconstituted suspension contained 250 mg of a polymer composition based on proanthocyanidin.

The invention described and claimed in this application is not limited to the specific embodiments proposed in this application. Embodiments of the invention are intended as examples of several aspects and embodiments. Any equivalent embodiments are within the scope of the present invention. Various variations of the invention in addition to those presented and described in this application will become apparent to those skilled in the art from the foregoing description. These options also fall under the attached claims. All publications cited in this application are incorporated into this description by reference in their entirety.

Claims (5)

CLAIM 1. A method of obtaining a polymer composition based on proanthocyanidin, including: a) providing a solution of milky juice obtained from the plant Csoyup 1esNeg1. containing sediment; b) adding methyl ethyl ketone to the plant’s milky juice solution and sediment; c) separating the organic layer and concentrating the aqueous layer to obtain a solid; or separating the aqueous layer and concentrating the organic layer to obtain a solid; b) dissolving the solid in an aqueous solvent; e) chromatography of the solution using an eluting solvent; and ί) isolation of proanthocyanidin containing Crofelemer. 2. The method according to claim 1, characterized in that step (e) of the chromatography is carried out by single-column or double-column chromatography on columns selected from CM-8erococcus Cancer! P1ote CoItp and 8erbabeh H-20. 3. The method according to claim 1, characterized in that the chromatography in step (e) is carried out using single-column chromatography on a column of SM-Sperox cancer! P1o \\\ 4. The method according to claim 1, characterized in that the chromatography in step (e) is carried out using double column chromatography on a SM-8erBagoke Cancer column! P1o \ y and the following ZerBabeh LN-20. 5. The method according to claim 1, characterized in that the chromatography in step (e) is carried out using water and then a 30% solution of acetone in water as eluting solvents. The result of an AKA sample of a partially purified polymer composition before treatment with bentonite in an aqueous solution Title time retention Square Conc. one nmol Conc. one TG 195 56971 0000000 0.000000 2.60 3777 0.000000 0.000000 3.34 5481 0.000000 0.000000 479 3316 0.000000 0.000000 aspartic acid 11 13 10503560 177059 233895 13.31 7765 0.000000 0.000000 19.54 580912 0.000000 0.000000 serine 17.21 7816924 12.0992 1271.63 21.55 1165044 0000000 0.000000 May 26 7220 0.000000 0.000000 glycine 34.32 30772317 44.1994 3319 38 alanine 36.25 11386972 17.8188 1587.65 39.76 864308 0.000000 0.000000 43.09 2460625 0.000000 0.000000 47.51 14475 0000000 0.000000 isoleucine 49.38 1113607 1.75237 229.910 leucine 50.81 21189965 30.7150 4029.60 phenylalanine 58.87 8073675 11.6630 1926.73 70.33 51138 0.000000 0000000 73.39 16083 0.000000 0.000000 74.19 91193 0000000 0.000000 76.33 2478 0.000000 0.000000 78.91 24748 0.000000 0.000000 84.05 12886267 0.000000 0.000000 97.01 22666 0.000000 0.000000 LYSINE 100.37 210701 0.291006 42.5451 histidine 105.92 211967 0.305738 47.4505 111.86 3593 0.000000 0.000000 115.39 18718 0000000 0.000000 arginine 131.90 185273 0.264437 46.0633 125 37 3121 0.000000 0.000000 125.93 1293 0.000000 0.000000 136.69 273299 0.000000 0.000000 1.098E + 08 136.815 14840.1 Peak reject level: 3000 FIG. one - 31 023814 The result of an AKA sample of a partially purified polymer composition after treatment with bentonite in an aqueous solution Title time retention Square Conc. one nmol Conc. one Ye aspartic acid 11.02 5333936 8.40385 1110.15 threonine 15 43 430151 0.606634 72.2501 serine 05.17 3974868 5.92141 622.340 glutamic acid 21.37 797633 1 29503 190.499 glycine 34.21 14799067 200148 1503.11 alanine 36.10 5195324 7.90740 709.440 valine 42.48 532892 0.907525 106.362 isoleucine 49 18 7351090 1.11568 146.377 leucine 50.72 10216076 137809 1808.03 tyrosine 54.09 30292 0.0404603 733301 phenylalanine 58.91 4159026 5.71465 944.060 lysine 100.75 134627 0.100216 26.3476 histidine 106.26 99968 0.134710 20.9071 arginine 122.15 104406 0.152230 26.5185 96469369 66.1355 7289.72 Peak reject level: 1000 FIG. 2 Increased amount of product after treatment with bentonite,