Highly purified and / or modified Fukan composition for the treatment of fibrous adhesions

Abstract

To provide a highly purified and / or modified fucan composition for treating fibrous adhesions.SOLUTION: There are provided compositions comprising purified / modified fucans having specific levels of desired and specified fucan components, methods of obtaining purified / modified fucans, and methods of using compositions comprising purified / modified fucans. The fucan composition comprises at least 96% w / w of fucan and 4% w / w or less of non-fucan components or compounds or substances, wherein the fucan comprises (i) fucose and sulfate, and has (ii) 20% w / w to 60% w / w of sulfate level; and contains (iii) 10% w / w or more and less than 17% w / w of counterions, and may further be composed of galactose, glucose, rhamnose, mannose, xylose, and glucuronic acid.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] Claims of Priority This application claims the benefit of U.S. Provisional Application No. 62 / 711,364, filed Jul. 27, 2018; U.S. Provisional Application No. 62 / 711,372, filed Jul. 27, 2018; U.S. Provisional Application No. 62 / 711,335, filed Jul. 27, 2018; U.S. Provisional Application No. 62 / 713,399, filed Aug. 1, 2018; U.S. Provisional Application No. 62 / 722,135, filed Aug. 23, 2018; U.S. Provisional Application No. 62 / 755,311, filed Nov. 2, 2018; U.S. Provisional Application No. 62 / 793,514, filed Jan. 17, 2019; U.S. Provisional Application No. 62 / 861,223, filed Jun. 13, 2019; co-pending U.S. Provisional Application No. 62 / 713,392, filed Aug. 1, 2018; U.S. Provisional Application No. 62 / 713,413, filed Aug. 1, 2018; U.S. Provisional Application No. 62 / 722,137, filed Aug. 23, 2018; U.S. Provisional Application No. 62 / 755,318, filed Nov. 2, 2018; U.S. Provisional Application No. 62 / 861,228, filed Jun. 13, 2019; co-pending U.S. Provisional Application No. 62 / 755,328, filed Nov. 2, 2018; U.S. Provisional Application No. 62 / 793,654, filed Jan. 17, 2019; and U.S. Provisional Application No. 62 / 861,235, filed Jun. 13, 2019, the contents of which are incorporated herein by reference.

Background Art

[0002] Fucoidan (including fucan) is a sulfated polysaccharide. Generally speaking, this means that they are molecules composed of many saccharides and also have sulfur atoms attached to the saccharides. The main saccharides have six carbon atoms and the chemical formula C6H 12The sugar containing O5 is called "fucose." "Fucoidan" (or fucoidin) refers to fucane derived from brown algae (seaweed). Fucan can exist alone or in the form of mixtures with other sugars, such as xylose, galactose, glucose, glucuronic acid, and / or mannose. These other sugars can be extracted from seaweed containing fucane or other raw materials. Fucan is currently derived from natural raw materials such as brown algae (seaweed) and sea cucumbers, but "fucan" as described herein includes polymer molecules having the chemical and structural motifs of fucane as considered herein, regardless of the (one or more) ultimate raw materials of fucane.

[0003] Fucoidan is Adenocystis utricularis, Ascophyllum nodosum, Chorda filum, Cystoseirabies marina, Durvillaea antarctica, Ecklonia kurome, Ecklonia maxima, Eisenia bicyclis, Fucus evanescens, Fucus vesiculosis, Hizikia fusiforme, Himanthalia Elongata, Kjellmaniella crassifolia, Laminaria brasiliensis, Laminaria cichorioides, Laminaria hyperborea, Laminaria japonica, Laminaria saccharina, Lessonia trabeculata, Macrocystis pyrifera, Pelvetia fastigiata, Pelvetia canaliculata, Saccharina japonica, Saccharina latissima, Sargassum stenophylum, Sargassum thunbergii, Sargassum These examples can be derived from a diverse range of brown algae species, including, but are not limited to, confusum, Sargassum fusiforme, and Undaria pinnatifida. All of these exemplary species belong to the taxonomic class Phaeophyceae, and the majority of these species belong to the families Fucales or Laminariaceae. [Overview of the Initiative] [Problems that the invention aims to solve]

[0004] Fukan, containing fucoidan, has been shown to be effective as a barrier device for preventing, inhibiting, or treating the formation of fibrous adhesions. Fukan has also been found to have applications in the treatment of other related diseases and conditions. Therefore, there is still an unmet demand for purified fucan preparations containing fucan modified to have a desired molecular weight distribution and / or sulfation level. This composition, system, and method, etc., offer these and / or other advantages. [Means for solving the problem]

[0005] Compositions, methods, systems, etc., are provided for fukan and fukan-containing compositions to suppress fibrous adhesions, among other advantages. Fukan and fukan compositions as used herein include fukan containing purified / modified fukan having a specific level of a desired identified fukan component. Purified / modified fukan and purified / modified fukan compositions are frequently referred to in the text of this application. Such references include all fukan / fukan compositions described herein, except as in the claims or unless explicitly limited to purified / modified fukan and / or purified / modified fukan compositions from the context. In some embodiments, the fukan and compositions containing them are suitable for medical and surgical applications. The fukan and fukan compositions have reduced levels of non-fukan components or impurities found in the raw fukan compositions. Such undesirable components or impurities include, for example, undesirable components that are bound to fukan (e.g., ionic, covalent, hydrogen, etc.), as well as compounds in the composition that are not part of fukan or are not chemically and / or ionically bound to fukan. Undesirable fukan components can be quantified relative to the fukan (e.g., by mass (w / w)). Non-fukan compounds and undesirable fukan components may collectively be referred to as impurities of the fukan and / or fukan-containing compositions as used herein. These purified / modified fukan can, for example, reduce dangerous complications in the medical and surgical use of fukan caused by impurities.

[0006] The present composition, method, system, etc., provides a composition containing a desired fucan, including a purified / modified fucan obtained from a starting fucan composition or an initial fucan composition (i.e., a fucan composition from which the modified fucan may be derived; such starting fucan composition may be crude or not, and may be pre-treated, an example of which is a feedstock fucan composition), as well as a method for obtaining such desired purified / modified fucan and a method for using such composition.

[0007] In some embodiments, the systems, apparatus, methods, etc. described herein provide a composition containing fucane in which the total content of fucose, galactose, sulfate, and counterions is greater than 90% w / w, greater than 92% w / w, greater than 94% w / w, greater than 95% w / w, greater than 97% w / w, greater than 97.5% w / w, greater than 98% w / w, greater than 98.8% w / w, greater than 99% w / w, greater than 99.5% w / w, or greater than 99.9% w / w. In some embodiments, the fucose content may be greater than 25% w / w, greater than 30% w / w, or greater than 35% w / w, and the galactose content may be less than 10% w / w or less than 5% w / w. The counterion content may be less than 17% or less than 14%. The counterions may be pharmaceutically acceptable counterions such as aluminum, arginine, benzathine, chloroprocaine, choline, sodium, potassium, lithium, ammonium, ethylenediamine, diethylamine, diethanolamine, ethanolamine, histidine, lysine, N-methylglucamine, meglumine, procaine, triethylamine, zinc, calcium, and magnesium. In some embodiments, the pharmaceutically acceptable counterions may contain, consist of, or essentially consist of at least one of sodium and potassium.

[0008] In certain embodiments, the composition containing the purified / modified fucane provided may have a total content of fucose, galactose, and sulfuric acid greater than 75% w / w, greater than 80% w / w, or greater than 84% w / w. The molecular weight distribution may have more than 60% w / w of the distribution greater than 100 kDa, the weight-average molecular weight may be greater than 100 kDa, and / or the peak molecular weight may be 70 kDa. An example of an approach for measuring such a distribution is: One 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm and packed with a hydroxylated polymethacrylate gel with an effective molecular weight range of approximately 50 kDa to approximately 5,000 kDa, another 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm and packed with a hydroxylated polymethacrylate gel with an effective molecular weight range of approximately 1 kDa to approximately 6,000 kDa, and a 40 mm guard column with an inner diameter of 6 mm and packed with a hydroxylated polymethacrylate gel, wherein the two analytical gel permeation chromatography columns and the one guard column are housed in a column compartment at approximately 30°C; A differential refractive index detector with a temperature of approximately 30°C; 0.6 mL / min 0.1 M sodium nitrate mobile phase flow; and Quantification of the peak molecular weight standard curve, which is essentially composed of the following dextran standards: 1st dextran standard with a peak molecular weight of approximately 2,200 kDa, 2nd dextran standard with a peak molecular weight of approximately 720 kDa to 760 kDa, 3rd dextran standard with a peak molecular weight of approximately 470 kDa to 510 kDa, 4th dextran standard with a peak molecular weight of approximately 370 kDa to 410 kDa, 5th dextran standard with a peak molecular weight of approximately 180 kDa to 220 kDa, and 6th dextran standard with a peak molecular weight of approximately 40 kDa to 55 kDa. This involves using a substantially aqueous gel permeation chromatography configuration. The peak molecular weight standard curve may further include dextran standards with peak molecular weights of approximately 3 kDa to 5 kDa.

[0009] The provided composition containing Fucan may have a number average molecular weight greater than 50 kDa and a sulfation level of about 20% w / w to 60% w / w, about 30% w / w to 55% w / w, or about 35% w / w to 52% w / w.

[0010] The total carbohydrate content may be between 27% w / w and 80% w / w. The total fucose content as a percentage of the total carbohydrate content may be approximately 30% w / w or more, approximately 50% w / w or more, approximately 70% w / w or more, approximately 90% w / w or more, or approximately 95% w / w or more. The total galactose content as a percentage of the total carbohydrate content may be less than approximately 60% w / w or less than approximately 20% w / w. The total content of glucuronic acid, mannose, rhamnose, glucose, and xylose as a percentage of the total carbohydrate content may be less than approximately 30% w / w.

[0011] Furthermore, this specification includes methods for producing or using the purified / modified Fukan described herein. Such use includes treating fibrous adhesions.

[0012] In some embodiments, the Fukan composition described herein comprises a solid Fukan composition containing 90% w / w or more of Fukan, such as the purified / modified Fukan described in the specification, and the solid Fukan composition may further be a solid composition containing a water content of less than 7% w / w, less than 6% w / w, less than 5% w / w, less than 4% w / w, less than 3% w / w, less than 2% w / w, less than 1% w / w, or less than 0.1% w / w.

[0013] In certain embodiments, the composition described herein is a medically acceptable composition comprising a therapeutically effective amount of the purified / modified fucan described herein in a medically acceptable buffer or diluent. The method described herein comprises treating fibrous adhesions in an animal such as a human, comprising selecting the purified / modified fucan described herein to suppress fibrous adhesions and administering the composition described herein, or the composition containing fucan in a dose range of 0.5 mg / kg to 50 mg / kg in the composition, to the injured site of the animal. The dose range may be 1 mg / kg to 25 mg / kg. The method may also involve the use of dose ranges including 0.5 mg / kg to 50 mg / kg or 1 mg / kg to 25 mg / kg to treat a target disease or disorder, for example, fibrous adhesions.

[0014] Furthermore, in order to prepare a composition containing purified / modified fukan, a method for preparing a composition containing purified / modified fukan described herein, for example, a method for removing impurities from a starting fukan composition such as a raw fukan composition, is provided herein. Such a method is, for example, To provide a starting Fukan composition containing impurities, A reaction mixture is generated by adding a flocculant to the starting Fukan composition. The impurities are agglomerated by heating the reaction mixture to produce agglomerated impurities, and This includes removing the aggregated impurities.

[0015] Providing the starting fukan composition may include providing the starting fukan composition as a solution, and the method may further include recovering the purified / modified fukan in a solution from which impurities have been reduced. Coagulating the impurities may include heating the reaction mixture at a pressure exceeding atmospheric pressure, and the coagulation aid may include salts such as alkali metals, alkaline earth metals, aluminum, and / or ammonium chlorides, bromides, iodides, fluorides, sulfates, sulfites, carbonates, bicarbonates, phosphates, nitrates, nitrites, acetates, citrates, silicates, and / or cyanides. The coagulation aid may also include bases such as alkali metals, alkaline earth metals, aluminum, and / or ammonium hydroxides and / or oxides.

[0016] The impurities removed may include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0017] A further method for removing impurities from a starting fukan composition to obtain purified / modified fukan is: To provide a starting Fukan composition that is a solid, and an extraction medium configured to dissolve impurities but unable to dissolve Fukan, The process involves mixing the starting Fukan composition with the extraction medium to produce a mixture of purified / modified Fukan and the extraction medium, and This may include separating the purified / modified Fukan from the extraction medium.

[0018] The method may further include recovering a composition comprising the purified / modified fucoidan that is solid. The extraction medium may include at least one organic solvent having a relative polarity of less than 0.765. The value of the relative polarity can be normalized from the measured value of the solvent shift of the absorption spectrum. See, for example, Christian Reichardt, Solvents and Solvent Effects in Organic Chemistry, Wiley-VCH Publishers, 3rd ed., 2003. The organic solvent may include at least one of ethanol, isopropanol, methanol, benzene, diethyl ether, decamethylcyclopentasiloxane, ethyl acetate, butanol, hexane, heptane, heptanol, octanol, and decanol. The extraction medium may further include at least one of a base, a surfactant, and an oxidizing agent. Providing the solid starting fucoidan composition may include precipitating the starting fucoidan composition from a solution. The impurities to be removed may include at least one of particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cell components, and DNA.

[0019] A further method for removing impurities from a starting fucoidan composition to obtain a purified / modified fucoidan may comprise providing a starting fucoidan composition containing impurities including suspended impurities in a solution, forming a mixture of suspended impurities, precipitated impurities, and a supernatant solution by precipitating the impurities from the solution using an ionic polyvalent impurity precipitant, and separating the suspended impurities and the precipitated impurities from the supernatant solution. The method may further include recovering the supernatant solution comprising the composition comprising the purified / modified fucoidan.

[0020] The ionic polyvalent impurity precipitant may contain a salt of a divalent or trivalent cation. The salt may be chloride, bromide, iodide, fluoride, sulfate, sulfite, carbonate, bicarbonate, phosphate, nitrate, nitrite, acetate, citrate, silicate, and / or cyanide. The cation may be an alkaline earth metal, zinc, aluminum, copper, and / or iron. The ionic polyvalent impurity precipitant may contain a base of a divalent or trivalent cation. The base may be a hydroxide and / or an oxide of an alkaline earth metal, zinc, aluminum, copper, and / or iron.

[0021] Separating the suspended impurities and the precipitated impurities from the supernatant solution may include aggregating the suspended impurities and the precipitated impurities by adding a flocculant to a mixture of the suspended impurities, the precipitated impurities, and the supernatant solution. The flocculant may include at least one of potassium aluminum sulfate, sodium aluminum sulfate, ammonium aluminum sulfate, calcium chloride, sodium phosphate, aluminum hydroxide, aluminum chloride, ferric chloride, ferric sulfate, ferrous sulfate, sodium silicate, calcium silicate, calcium phosphate, zinc chloride, calcium carbonate, calcium bicarbonate, potassium sulfate, magnesium phosphate, acrylamide, acrylic acid, aluminum chlorohydrate, polyaluminum chloride, tannin, formaldehyde, melamine, N,N-dimethylaminoethyl acrylate methyl chloride, N,N-dimethylaminoethyl methacrylate methyl chloride quaternary salt, and polydiallyldimethylammonium chloride. The method may further include maintaining a pH of about 7 to 14. Maintaining the pH may include adding a base. The impurities to be removed may include at least one of particulate matter, lipid, fatty acid, phlorotannin, laminarin, alginate, protein, Maillard reaction product, fucoxanthin, chlorophyll, bacteria, cell components, and DNA.

[0022] A method for removing impurities from a starting fucoidan composition to obtain a purified / modified fucoidan is providing a starting fucoidan composition containing impurities, Adjust the pH of the aforementioned starting Fukan composition to approximately 8-14. Adding a cell-disrupting agent configured to dissolve cellular components to the starting Fukan composition to produce a reaction mixture containing the cell-disrupting agent, a biomolecular lysate, and the starting Fukan composition, and This may include removing the cell-destroying agent and biomolecular lysate from the reaction mixture.

[0023] Providing the starting Fukan composition may include providing the starting Fukan composition as a solution, and the method may further include recovering the composition containing purified / modified Fukan in a solution with reduced impurities. The cell disruptor may include a surfactant which may be an anionic surfactant, a cationic surfactant, or a nonionic surfactant. The surfactant may include at least one of sodium dodecyl sulfate (SDS), benzalkonium chloride, Triton X100®, Triton X114®, Brij® surfactant, Tween® surfactant, sodium deoxycholate, and alkylbenzene sulfonates. Removing the cell disruptor and the biomolecular lysate may include adding a flocculant configured to aggregate the cell disruptor and biomolecular lysate to the reaction mixture. Removing the cell disruptor may include adding a precipitating agent configured to make the cell disruptor insoluble in the reaction mixture to the reaction mixture to produce a precipitate. Removing the biomolecular lysate may include adding a precipitating agent configured to make the biomolecular lysate insoluble in the reaction mixture to the reaction mixture to produce a precipitate. The method may further include adding a flocculant configured to floccate the precipitate to the reaction mixture. The flocculant may include at least one of potassium aluminum sulfate, sodium aluminum sulfate, ammonium aluminum sulfate, calcium chloride, sodium phosphate, aluminum hydroxide, aluminum chloride, ferric chloride, ferric sulfate, ferrous sulfate, sodium silicate, calcium silicate, calcium phosphate, zinc chloride, calcium carbonate, calcium bicarbonate, potassium sulfate, magnesium phosphate, acrylamide, acrylic acid, aluminum chlorohydrate, polyaluminum chloride, tannin, formaldehyde, melamine, N,N-dimethylaminoethyl acrylate methyl chloride, N,N-dimethylaminoethyl methacrylate methyl chloride quaternary salt, and polydiallyldimethylammonium chloride.

[0024] The removal of the anionic surfactant may include anionic adsorption. The removal of the cationic surfactant may include cationic adsorption. The removal of the nonionic surfactant may include micelle phase separation. Furthermore, the removal of the surfactant may include hydrophobic adsorption. The removal of the surfactant may include, Diluting the reaction mixture until the concentration of the surfactant falls below a predetermined concentration, and The reaction mixture containing the surfactant may be subjected to dialysis filtration using a tangential flow filter, the fractional molecular weight of which exceeds the maximum molecular weight of the surfactant.

[0025] The method may also include adding a chelating agent to the reaction mixture after providing the starting Fukan composition and before removing the cell-destroying agent. The chelating agent may include ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin, and / or citric acid. The method may further include adding an oxidant-quenching agent to the reaction mixture before removing the cell-destroying agent to quench the reaction of the oxidant in the reaction mixture, or adding a bacteriostatic agent to the reaction mixture after providing the starting Fukan composition and before removing the cell-destroying agent. The bacteriostatic agent may include sodium sulfite, ethylenediaminetetraacetic acid (EDTA), benzalkonium chloride, ethanol, and / or thiourea. The impurities removed may include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0026] A method for removing impurities from a starting fukan composition to obtain the composition containing purified / modified fukan is: To provide a starting Fukan composition containing impurities in an aqueous starting solution, The aqueous starting solution is mixed with an organic solvent to produce an aqueous-organic phase mixture, and The process may include separating the aqueous-organic-phase mixture to obtain an aqueous portion and an organic portion.

[0027] The method may further include recovering the aqueous portion containing the purified / modified fucan. The organic solvent may include at least one organic solvent with a relative polarity of less than 0.765, which may be at least one of ethanol, isopropanol, methanol, benzene, decamethylcyclopentasiloxane, ethyl acetate, hexane, heptanol, octanol, decanol, heptane, isobutyl acetate, anisole, isopropyl acetate, 1-butanol, butyl acetate, methyl isobutyl ketone, pentane, 1-pentanol, ethyl ether, and propyl acetate. The impurities removed may include at least one of particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0028] A method for changing the cation content of the starting Fukan composition is: To provide the starting Fukan composition in the starting solution, and The process may also include subjecting the starting solution to dialysis filtration through a tangential flow filtration filter using a chelating agent solution that has passed through a tangential flow filtration filter to produce a retained Fukan composition.

[0029] The chelating agent may comprise at least one of ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin, or citric acid. The retaining fukan composition may have a cation content consisting essentially of sodium and / or potassium.

[0030] A method for removing impurities from a starting fukan composition to obtain purified / modified fukan is further described as follows: To provide a starting Fukan composition containing impurities, The starting Fukan composition is subjected to an appropriate pressure exceeding 70 bar and an appropriate temperature exceeding 30°C in a supercritical extraction apparatus. The supercritical fluid is filled into a supercritical extraction apparatus to remove impurities from the supercritical fluid, and This may include removing the supercritical fluid containing the extracted impurities after a predetermined time.

[0031] The method may further include recovering the purified / modified fukan remaining in the supercritical extractor, the pressure may be about 70 bar to about 2000 bar, and the temperature may be about 30°C to about 300°C. The starting fukan composition may be liquid or solid. The supercritical fluid may contain at least one of carbon dioxide, ethanol, ethane, hydrochloric acid, hydrofluoric acid, sulfuric acid, and nitric acid. The predetermined time may be about 5 minutes to 50 hours. The impurities removed may contain at least one of particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0032] These and other embodiments, features, and models are shown in this application, including embodiments and accompanying drawings for carrying out the following inventions. Unless otherwise expressly stated, all embodiments, features, etc., can be harmonized and adapted, combined and rearranged in any desirable manner.

[0033] These and other embodiments, features, and models are shown in this application, including embodiments and accompanying drawings for carrying out the following inventions. Unless otherwise expressly stated, all embodiments, features, etc., can be harmonized and adapted, combined and rearranged in any desirable manner. [Brief explanation of the drawing]

[0034] [Figure 1]Figure 1 schematically illustrates an exemplary system for modifying the cation content of a starting Fucan composition and removing low molecular weight non-Fuccan components using a chelating agent in tangential flow filtration. [Figure 2A] NMR results are shown demonstrating that certain types of Fucans treated according to the methods described herein undergo structural changes. [Figure 2B] Two-dimensional NMR results are shown demonstrating that certain Fukans treated according to the methods described herein undergo structural changes. [Modes for carrying out the invention]

[0035] The aforementioned drawings illustrate exemplary embodiments of several aspects of the compositions, methods, etc., described herein. Embodiments of systems, methods, etc., described herein may include further features or steps not shown in the drawings. Furthermore, the examples described herein illustrate embodiments of systems, methods, etc., in one or more forms, and such examples should not be construed as limiting the scope of the disclosure in any way. The embodiments described herein are not exhaustive and the disclosure is not limited to the detailed forms disclosed, for example, in the embodiments for carrying out the following inventions.

[0036] The compositions, systems, methods, etc., described herein include purified / modified fucoidan. The compositions herein may be effective for medical procedures, post-operative care, disease control, etc. In some embodiments, the fucoidan is fucoidan. The purified / modified fucoidan herein may be medical devices, medical materials, combination products, etc., may be included therein, or may be included in pharmaceutically acceptable therapeutic and / or medically effective compositions.

[0037] The following paragraphs move on to a brief and general discussion of some of the compositions described herein, including purified / modified Fucan. These compositions include those that can be prepared using the methods described herein from a starting Fucan composition in a variety of ways, depending on the method chosen, using a suitable reaction mixture such as a solution, suspension, solid, gel, or other modality.

[0038] composition The compositions, systems, etc., described herein provide, in certain embodiments, phytoacceptable purified / modified fucane compositions comprising fucane and phytoacceptable purified / modified fucane, as well as therapeutically effective amounts of the purified / modified fucane composition for treating fibrous adhesions in arthritis, psoriasis, or optionally other diseases, as well as surgical adhesions. The purified / modified fucane in the composition may have a total content of fucose, galactose, and sulfate of more than about 75% w / w, for example, more than about 80% w / w or more than 84% w / w. In some embodiments, the purified / modified fucane may further contain at least one counterion up to at least about 5% w / w, 7% w / w, 9% w / w, 10% w / w, or 11% w / w. In some embodiments, the counterion is a phytoacceptable counterion. In some embodiments, the counterion is ionically bonded to a sulfate group present on the fucane. pharmaceutically acceptable counterions may include aluminum, arginine, benzathine, chloroprocaine, choline, sodium, potassium, lithium, ammonium, ethylenediamine, diethylamine, diethanolamine, ethanolamine, histidine, lysine, N-methylglucamine, meglumine, procaine, triethylamine, zinc, calcium, and magnesium. The sulfur-containing components of Fucan are linked via COS bonds. The oxygen in such bonds can be considered to be primarily bonded to either carbon or sulfur, depending on various factors. As used herein, the term “sulfuric acid” refers to both embodiments.

[0039] In certain embodiments, the purified / modified fucane described herein contains at least about 85% w / w, 90% w / w, 94% w / w, 97% w / w, or 98% w / w of fucose, galactose, sulfate, and counterions. Exemplary counterions include up to about 7% w / w, 8% w / w, 9% w / w, 10% w / w, 11% w / w, 12% w / w, 13% w / w, 14% w / w, or 15% w / w of calcium, magnesium, potassium, and / or sodium. In some embodiments, the purified / modified fucane contains at least about 25% w / w, 30% w / w, or 35% w / w of fucose. In some embodiments, the purified / modified fucane contains about 10% w / w, 5% w / w, or less than 4% w / w of galactose. In some embodiments, the purified / modified fucane consists essentially of the sum of fucose, galactose, sulfate, and counterionic components, or consists of the sum of fucose, galactose, sulfate, and counterionic components. In some embodiments, the fucane described herein is substantially or completely lacking in all sugar components other than fucose and galactose. In some embodiments, the fucane described herein is substantially or completely lacking in one or more of glucuronic acid, mannose, rhamnose, xylose, galactose, or glucose. As used in this sentence, “substantially lacking” means that even if such sugar components are present, their presence is so low that it is not pharmaceutically or medically significant.

[0040] In certain embodiments, the purified / modified Fukan described herein may be used for a number of applications including the suppression, prevention, removal, mitigation, or other treatment of fibrous adhesions and other targets such as other diseases and / or conditions. Treatments include reducing or preventing the manifestation of a target disease or other condition, such as Fukan reducing or preventing the formation of fibrous adhesions at a target site which is a selective target site identified by a surgeon or other specialist as containing or being considerably susceptible to fibrous adhesions (or other diseases or conditions), and also including the removal of existing diseases or other conditions, such as the removal of already existing fibrous adhesions. For such suppression, prevention, removal, mitigation, or other treatment, the Fukan composition is typically provided in a pharmaceutically effective composition comprising additional components such as medically acceptable medical devices, combination products, or binders, adjuvants, excipients, and optionally additional medically active substances, such as secondary drugs that are not added to the Fukan but are included in the composition and / or can be added to the Fukan.

[0041] In further embodiments, the composition comprising purified / modified Fucan described herein may be a solid, for example, a solid composition having a water content of less than about 7% w / w, for example, less than about 6% w / w, less than 5% w / w, less than 4% w / w, less than 3% w / w, or less than 2% w / w.

[0042] The molecular weight distribution of the purified / modified Fucan can be measured using any desired suitable measurement system. Different systems may produce different readings or results from different compositions having substantially the same composition, or from the same batch if measured differently. One suitable measurement system comprises: one 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm, packed with a hydroxylated polymethacrylate gel with a substantially effective molecular weight range of about 50 kDa to about 5,000 kDa; one 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm, packed with a hydroxylated polymethacrylate gel with a substantially effective molecular weight range of about 1 kDa to about 6,000 kDa; and one 40 mm guard column with an inner diameter of 6 mm, packed with a hydroxylated polymethacrylate gel, all housed in a column compartment at about 30°C. The aqueous gel permeation chromatography configuration consists of a guard column, a differential refractive index detector at approximately 30°C, a 0.1 M sodium nitrate mobile phase flow of 0.6 mL / min, and quantification of a peak molecular weight standard curve consisting substantially of a first dextran standard with a peak molecular weight of approximately 2,200 kDa, a second dextran standard with a peak molecular weight of approximately 720 kDa to 760 kDa, a third dextran standard with a peak molecular weight of approximately 470 kDa to 510 kDa, a fourth dextran standard with a peak molecular weight of approximately 370 kDa to 410 kDa, a fifth dextran standard with a peak molecular weight of approximately 180 kDa to 220 kDa, and a sixth dextran standard with a peak molecular weight of approximately 40 kDa to 55 kDa. The peak molecular weight standard curve may further include dextran standards with peak molecular weights of 3 kDa to 5 kDa.

[0043] The purified / modified Fucan described herein may have a molecular weight distribution, where at least about 25% w / w, 30% w / w, 40% w / w, 50% w / w, 60% w / w, 70% w / w, 75% w / w, 90% w / w, 92% w / w, 97% w / w, or 98% w / w of the distribution is greater than 100 kDa. The purified / modified Fucan described herein may contain Fucan having a molecular weight distribution, where at least about 50% w / w, 60% w / w, 70% w / w, 80% w / w, or 90% w / w of the distribution is greater than 200 kDa. The purified / modified Fucan described herein may have a molecular weight distribution, wherein at least about 25% w / w, 30% w / w, 40% w / w, 50% w / w, 60% w / w, 70% w / w, or 75% w / w of the distribution is greater than 500 kDa. The purified / modified Fucan described herein may have a molecular weight distribution, wherein at least about 5% w / w, 10% w / w, 20% w / w, 30% w / w, or 40% w / w of the distribution is greater than 1600 kDa.

[0044] The purified / modified fukan described herein may have a weight-average molecular weight greater than approximately 100 kDa, for example, approximately 100 kDa to approximately 10,000 kDa, approximately 200 kDa to approximately 8,000 kDa, approximately 350 kDa to approximately 8,000 kDa, approximately 450 kDa to approximately 8,000 kDa, approximately 580 kDa to approximately 8,000 kDa, or approximately 800 kDa to approximately 2,000 kDa. The purified / modified fucan described herein may have a peak molecular weight greater than approximately 70 kDa, for example, approximately 70 kDa to approximately 1200 kDa, approximately 100 kDa to approximately 1200 kDa, approximately 200 kDa to approximately 1200 kDa, approximately 400 kDa to approximately 1200 kDa, or approximately 400 kDa to approximately 900 kDa.

[0045] The purified / modified Fucan described herein may have a number average molecular weight of more than approximately 50 kDa, approximately 50 kDa to approximately 1,000 kDa, approximately 70 kDa to approximately 1,000 kDa, approximately 150 kDa to approximately 1,000 kDa, approximately 250 kDa to approximately 1,000 kDa, or approximately 250 kDa to approximately 700 kDa.

[0046] The purified / modified Fucan described herein may have a sulfated level of approximately 10% w / w to 70% w / w, approximately 20% w / w to 65% w / w, approximately 30% w / w to 60% w / w, or approximately 40% w / w to 60% w / w.

[0047] The purified / modified fucane described herein may have a total fucose:total sulfuric acid molar ratio of approximately 1:0.5 to 1:4, approximately 1:0.8 to 1:3.5, approximately 1:1 to 1:2.5, approximately 1:1.2 to 1:2.0, or approximately 1:1.5 to 1:3. The purified / modified fucane described herein may have a total fucose + galactose:total sulfuric acid molar ratio of approximately 1:0.5 to 1:4, approximately 1:0.8 to 1:3.5, approximately 1:1 to 1:2.5, approximately 1:1.2 to 1:2.0, or approximately 1:1.5 to 1:3.

[0048] The purified / modified fucane described herein may have a total carbohydrate content of approximately 27% w / w to 70% w / w, approximately 30% w / w to 80% w / w, approximately 40% w / w to 90% w / w, or approximately 50% w / w to 100% w / w. The purified / modified fucane described herein may have a fucose content of approximately 30% w / w to 100% w / w, approximately 40% w / w to 95% w / w, or approximately 50% w / w to 90% w / w as a percentage of the total carbohydrate content. The fucane described herein may have a galactose content of 0% w / w to 60% w / w, approximately 5% w / w to 30% w / w, or approximately 8% w / w to 10% w / w as a percentage of the total carbohydrate content. The Fucan described herein may have a glucuronic acid content of approximately 0% w / w to 10% w / w relative to the total carbohydrate content, a mannose content of approximately 0% w / w to 7% w / w relative to the total carbohydrate content, a rhamnose content of 0% w / w to 4% w / w relative to the total carbohydrate content, and a xylose content of 0% w / w to 20% w / w relative to the total carbohydrate content. The Fucan described herein may have a total content of glucuronic acid, mannose, rhamnose, glucose, and xylose of less than approximately 30% w / w or less than approximately 12% w / w.

[0049] In some embodiments, the purified / modified fukan described herein, when dissolved in water at a concentration of 50 mg / ml, has viscosities of about 4 cP to about 50 cP, about 5 cP to about 40 cP, about 10 cP to about 30 cP, about 15 cP, about 20 cP, or about 25 cP. In certain embodiments, when the purified / modified fukan described herein is dissolved in water at a concentration of 1 mg / ml to 100 mg / ml, it forms a solution that is colorless, pale yellow, or pale brown.

[0050] The purified / modified Fukan described herein may be provided in the form of a paste, gel, patch, film, spray, liquid, lotion, cream, solution, suspension, solid, implant, microsphere, or other desired form.

[0051] The compositions described herein may be solids consisting essentially of purified / modified fucane. Purified / modified fucane may consist essentially of fucose, galactose, sulfuric acid, and counterions.

[0052] In this specification, purified / modified fucane may be present in solutions containing fucane at concentrations of about 0.01 mg / mL to about 300 mg / mL, for example, about 0.1 mg / mL to about 100 mg / mL, about 1 mg / mL to about 50 mg / mL, or about 20 mg / mL to about 80 mg / mL. The fucane may essentially consist of fucose, galactose, sulfate, and counterions.

[0053] The purified / modified fucane may be present in a gel containing fucane at concentrations of approximately 100 mg / mL to approximately 1000 mg / mL, for example, approximately 100 mg / mL to approximately 500 mg / mL or 300 mg / mL to approximately 800 mg / mL. The fucane may essentially consist of fucose, galactose, sulfate, and counterions.

[0054] In this specification, purified / modified fucane may be present in films containing fucane at concentrations of approximately 100 mg / mL to approximately 1000 mg / mL, for example, approximately 100 mg / mL to approximately 500 mg / mL or approximately 300 mg / mL to approximately 800 mg / mL. The fucane may essentially consist of fucose, galactose, sulfate, and counterions.

[0055] The purified / modified Fucan described herein may be administered as components of medical devices, combination products, and / or pharmaceutical compositions comprising any number of pharmaceutically acceptable excipients, such as gelatin, hypromellose, lactose, water for injection (USP compliant), sodium chloride, sodium phosphate, sodium citrate, sodium ascorbate, phosphate buffer, citrate buffer, phosphate-citrate buffer, Pluronic acid, cellulose, alginate, acrylate, hyaluronic acid, polyethylene glycol, chitosan, excipients for injection, or Ringer's lactate solution USP.

[0056] The purified / modified Fucan described herein may be administered in the form of a paste, gel, patch, film, spray, liquid, lotion, cream, solution, suspension, solid, implant, microsphere, or other desired form.

[0057] The purified / modified Fucan may be administered intravenously, intraarticularly, intrafocally, vaginally, rectally, intramuscularly, intraperitoneally, subcutaneously, topically, intranasally, intraocularly, or orally. The purified / modified Fucan may be delivered directly to the affected area. The purified / modified Fucan may be continuously released to the affected area via controlled release from a polymer dosage form.

[0058] The purified / modified Fucan described herein may be administered as an ingredient in a pharmaceutical composition comprising the purified / modified Fucan and at least one other agent. The agent may be at least one of the following: paclitaxel, doxorubicin, camptothecin, etoposide, mitoxantrone, methotrexate, menadione, plumbagin, juglon, β-rapacon cyclosporine, sulfasalazine, steroids, rapamycin, retinoids, docetaxel, colchicine, antisense oligonucleotides, and ribozymes.

[0059] In certain embodiments, the purified / modified fucan described herein may have an acetyl content of less than about 5% w / w, less than about 2% w / w, or about 0% w / w. In some embodiments, the purified / modified fucan described herein may have an acetyl content of 10–30 ppm in the carbon dimension in eight increments of 256–512 scans, respectively, at 70°C with solvent signal suppression in a 600 MHz spectrometer equipped with a 5 mm cryogenic probe. 1 H- 13 When measured by C-henuclear multi-quantum coherence, the acetyl content is substantially 0% w / w.

[0060] method For example, methods and systems are provided for purifying and / or modifying fucane from a starting fucane composition containing fucane, such as a raw material fucane composition, or other fucane-containing compositions. As used herein, “impurities” means any component of fucane that is not fucose, galactose, sulfate, or counterions, and any non-fucan components, compounds, or substances present in a composition containing fucane. Such fucane may contain impurities such as proteins ionically and / or chemically bonded to fucane, sugar residues other than fucose and galactose that are part of the fucane polymer structure, other sugars chemically bonded to fucane, and non-fucan impurities that are not bonded to fucane but are present in starting fucane compositions such as raw material fucane compositions. Examples of such impurities include, but are not limited to, particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. Some of these include chromophores, and therefore the starting fucane composition may contain brown, yellow, and green colors, some of which may be ionic and / or chemically bonded to fucane or parts thereof in the starting fucane composition. In certain embodiments, purified / modified fucane can be prepared using methods such as those described herein, containing fucose, galactose, sulfate, and counterions in amounts of at least about 88% w / w, 89% w / w, 90% w / w, 91% w / w, 92% w / w, 93% w / w, 94% w / w, 95% w / w, 96% w / w, 97% w / w, 97.1% w / w, 98% w / w, 98.8% w / w, 99% w / w, 99.5% w / w, or 99.9% w / w. In some embodiments, the purified / modified fucane contains fucose, galactose, and sulfuric acid in amounts of at least about 75% w / w, 78% w / w, 80% w / w, 82% w / w, or 84% w / w. In some embodiments, the purified / modified fucane contains impurities in amounts of less than about 0.1% w / w, less than 0.5% w / w, less than 1% w / w, less than 2.9% w / w, less than 3% w / w, less than 4% w / w, less than 5% w / w, less than 6% w / w, less than 7% w / w, less than 8% w / w, less than 9% w / w, less than 10% w / w, less than 11% w / w, or less than 12% w / w.Some of these impurities may cause dangerous complications in the medical and / or surgical use of Fukan.

[0061] In some embodiments, the disclosure presents purified / modified Fucan with low levels of impurities suitable for medical and surgical applications, such as the prevention of fibrous adhesions.

[0062] The following paragraphs will move on to some brief and general descriptions of the methods that may be used to produce the purified / modified Fucan described herein.

[0063] Physically induced aggregation (flocculation) A starting fukan composition, such as a raw fukan composition containing high levels of impurities, is subjected to the aggregation of impurities, which may be physically induced. The above method may include: providing a starting fukan composition; adding an aggregation aid to the starting fukan composition to produce a reaction mixture; heating the reaction mixture to aggregate the impurities in the starting fukan composition; separating the aggregated impurities from the reaction mixture; and recovering the desired purified / modified fukan after the separation.

[0064] The aggregation of the impurities by heating the reaction mixture may include heating the reaction mixture while subjecting it to a pressure exceeding atmospheric pressure. Suitable flocculants include, but are not limited to, salts and / or bases, such as alkali metals, alkaline earth metals, aluminum and / or ammonium chlorides, bromides, iodides, fluorides, sulfates, sulfites, carbonates, bicarbonates, phosphates, nitrates, nitrites, acetates, citrates, silicates, oxides, hydroxides and / or cyanides, such as sodium chloride, sodium sulfate, potassium chloride, calcium sulfate, sodium phosphate, sodium nitrate, lithium chloride, lithium nitrate, ammonium chloride, sodium carbonate, and sodium hydroxide. Separation of the aggregated impurities from the reaction mixture may include one or more of centrifugation, filtration, precipitation, or hydrodynamic flow separation of the reaction mixture.

[0065] The methods described herein may further include desalting the starting Fukan composition before adding a flocculant. The desalting may include dialyseptic filtration of the starting Fukan composition, which is a solution in water, through a molecular weight cutoff (MWCO) tangential flow filtration (TFF) filter. The dialyseptic filtration may include dialyseptic filtration of the starting Fukan composition with distilled water. The molecular weight cutoff TFF filter may have a molecular weight cutoff smaller than the desired molecular weight separation point or target in the purified / modified Fukan, for example, 50 kDa, 70 kDa, 100 kDa, 200 kDa, 300 kDa, 500 kDa, or 1000 kDa.

[0066] The above method may be carried out in a basic or neutral environment. Adding a flocculant to the starting fukan composition may therefore include making the starting fukan composition basic to prevent or suppress the decomposition of fukan in the starting fukan composition, since fukan tends to decompose in an acidic environment. In other embodiments, the above method may be carried out by maintaining the reaction mixture at or near pH 7.

[0067] In some embodiments, the starting fucane composition may be provided as a solution. Examples of fucane suitable for treatment by the above method include, but are not limited to, fucoidan, and the fucane concentration in the solution may be 0.01% w / v to 50% w / v. Impurities that can be removed by the above method include, but are not limited to, particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0068] solid phase extraction For example, the fukan contained in a starting fukan composition, such as a raw fukan composition containing a very high level of impurities or an undesirable level of impurities, is subjected to solid-phase extraction. The method may include: providing a starting fukan composition in a solid state containing impurities among other impurities, and an extraction medium configured to dissolve impurities but unable to dissolve fukan; mixing the starting fukan composition with the extraction medium to form a mixture of an undissolved solid fukan composition and the extraction medium containing dissolved impurities; separating the purified, undissolved solid fukan from the extraction medium containing dissolved impurities; and recovering the purified / modified fukan as a solid after removing it from the extraction medium. The separation may include, for example, one or more of centrifugation, filtration, precipitation, and hydrodynamic fluid separation.

[0069] The extraction medium may include, for example, one or more of a base, a surfactant, and an oxidizing agent. Suitable extraction media that do not dissolve Fukan include organic solvents with a relative polarity of less than 0.765, such as ethanol, isopropanol, methanol, benzene, diethyl ether, decamethylcyclopentasiloxane, ethyl acetate, butanol, hexane, heptane, heptanol, octanol, and decanol. Suitable bases include, without limitation, sodium hydroxide, potassium hydroxide, lithium hydroxide, and calcium hydroxide. Suitable oxidizing agents include, without limitation, one or more of oxidizing bleaches, including hydrogen peroxide, urea peroxide, and sodium hypochlorite. Suitable surfactants include, without limitation, nonionic surfactants, such as those in the range of Tween®, Brij®, and Triton®; anionic surfactants, such as sodium dodecyl sulfate (SDS) and sodium deoxycholate; and cationic surfactants, such as benzalkonium chloride (BAC). Specific fucoidans suitable for the methods described herein include, but are not limited to, fucoidan. The treatment of the original, for example, starting fucoidan composition with the extraction medium can be extended from 1 minute to 120 hours.

[0070] The method may further include desalting the starting Fukan composition before providing the starting Fukan composition in solid form. Desalting may include dialyzing the starting Fukan composition, which is a solution in water, using a molecular weight cutoff (MWCO) tangential flow filtration (TFF) filter. Dialysis filtration may include dialyzing the starting Fukan composition with distilled water. The molecular weight cutoff TFF filter may have a molecular weight cutoff smaller than the desired molecular weight separation point or target in the purified / modified Fukan, for example, 50 kDa, 70 kDa, 100 kDa, 200 kDa, 300 kDa, 500 kDa, or 1000 kDa. Dialysis filtration may further include pre-filtration of the starting Fukan composition through a suitable pre-filter to remove particulate matter. The method may further include freeze-drying a suitable starting Fukan composition in solution before providing the starting Fukan composition in solid form. The method may further include precipitating a suitable starting Fukan composition from solution before providing the starting Fukan composition in solid form. Suitable precipitating agents include, but are not limited to, ethanol, isopropanol, propanol, acetone, methanol, dimethyl sulfoxide, dimethylformamide, ethylene glycol, tetrahydrofuran, acetonitrile, glyme, diglyme, and dioxane, and the solubility of fukan decreases as the polarity of the precipitating fluid decreases. Impurities that can be removed by the above method include, but are not limited to, particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0071] Chemically induced precipitation Starting fukan compositions, such as raw fukan compositions, containing high levels of impurities, for example, suspended particulate matter, are subjected to chemically derived impurity precipitation. In certain embodiments, the method may include: providing the starting fukan composition in a starting solution; precipitating the impurities from the starting solution using an ionic polyvalent impurity precipitant to provide a mixture of suspended impurities, precipitated impurities, and a supernatant; separating the suspended and precipitated impurities from the supernatant; and recovering the supernatant containing the desired purified / modified fukan after the suspension and precipitated impurities have been separated from the supernatant.

[0072] Suitable impurity precipitants include ionic polyvalent salts and / or bases of divalent and trivalent cations. Examples of such suitable salts, without limitation, include chlorides, bromides, iodides, fluorides, sulfates, sulfites, carbonates, bicarbonates, phosphates, nitrates, nitrites, acetates, citrates, silicates and / or cyanides of alkaline earth metals, zinc, aluminum, copper and / or iron. Examples of such suitable bases, without limitation, include hydroxides and / or oxides of alkaline earth metals, zinc, aluminum, copper and / or iron. Separation of suspended and precipitated impurities from the supernatant may involve agglomeration of impurities in the mixture. Suitable flocculants include, without limitation, potassium aluminum sulfate; sodium aluminum sulfate; aluminum ammonium sulfate; calcium chloride; sodium phosphate; aluminum hydroxide; aluminum chloride; ferric chloride; ferric sulfate; ferrous sulfate; sodium silicate; calcium silicate; calcium phosphate; zinc chloride; calcium carbonate; calcium bicarbonate; potassium sulfate; magnesium phosphate; acrylamide; acrylic acid; aluminum chlorohydrate; polyaluminum chloride; tannin; formaldehyde; melamine; N,N-dimethylaminoethyl acrylate methyl chloride; N,N-dimethylaminoethyl methacrylate methyl chloride quaternary salt; and polydiallyldimethylammonium chloride. As can be seen from the above enumeration of flocculants, in some embodiments the flocculant may be an impurity precipitating agent. Separation of precipitated, suspended and / or flocculated impurities from the supernatant may include at least one of centrifugation, filtration, precipitation, and hydrodynamic flow separation of the mixture of impurities and the supernatant.

[0073] The method may further include desalting the starting fukan composition before providing the starting fukan composition. The desalting may include dialyseptic filtration of the starting fukan composition as an aqueous solution through a TFF filter. The dialyseptic filtration may include dialyseptic filtration of the starting fukan composition with distilled water. The dialyseptic filtration may include dialyseptic filtration of the starting fukan composition through a TFF filter having an MWCO of 5 kDa, 10 kDa, 30 kDa, 50 kDa, 70 kDa, or 100 kDa. The dialyseptic filtration may further include pre-filtration of the starting fukan composition through a suitable pre-filter to remove particulate matter.

[0074] The method described above may further include maintaining a pH of about 7 to 14 to suppress or prevent the decomposition of fukan in an acidic environment. Maintaining this pH of about 7 to 14 may include the addition of a suitable base, such as sodium hydroxide. The suitable base may be added to the starting fukan composition before precipitating impurities from the solution using an ionic polyvalent impurity precipitating agent. In another embodiment, the suitable base may be added to a mixture of the precipitated impurities and the supernatant after precipitating impurities from the solution using an ionic polyvalent impurity precipitating agent. In yet another embodiment, the suitable base may be added to the supernatant after separating the suspended and precipitated impurities from the supernatant.

[0075] Examples of fucoidan suitable for treatment by the above method include, but are not limited to, fucoidan, and the fucoidan concentration in the solution can be 0.01% w / v to 50% w / v. Impurities that can be removed by the above method include, but are not limited to, particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0076] Dissolution and aggregation A starting fukan composition, such as a raw fukan composition containing high levels of impurities, is subjected to dissolution and aggregation. The method in this example may include: providing a starting fukan composition; making the starting fukan composition alkaline; adding a cell disruptor to the starting fukan composition to produce a reaction mixture, wherein the cell disruptor dissolves cellular components in the starting fukan composition and releases a lysate containing biomolecular components into an alkaline reaction mixture; and removing at least a portion of the cell disruptor and impurities from the reaction mixture to leave the desired fukan that has not been decomposed.

[0077] The removal of the cell-disrupting agent may include one or more of the following: precipitation, flocculation, tangential flow filtration, micelle phase separation, ion adsorption, and hydrophobic adsorption. The removal of impurities may include one or more of the following: precipitation, flocculation, tangential flow filtration, micelle phase separation, ion adsorption, and hydrophobic adsorption. Any or a combination of these removal methods may include centrifugation, filtration, precipitation, or hydrodynamic flow separation of a mixture of either the solid phase or the liquid phase.

[0078] Suitable cell disruptors include, but are not limited to, anionic, nonionic, or cationic surfactants, such as sodium dodecyl sulfate (SDS), benzalkonium chloride, Triton X100®, Triton X114®, Brij® surfactants, Tween® surfactants, sodium deoxycholate, and alkylbenzene sulfonates.

[0079] In one embodiment of the above method, the cell-disrupting agent is sodium dodecyl sulfate (SDS), and removal of the cell-disrupting agent includes adding a precipitating agent to precipitate the cell-disrupting agent by making it insoluble in the alkaline reaction mixture. In this embodiment, removal of the cell-disrupting agent may further include adding a flocculant to the reaction mixture to flocce the precipitated cell-disrupting agent together with at least some impurities. Removal of the cell-disrupting agent may further include centrifugation after flocculation.

[0080] Suitable precipitating agents for sodium dodecyl sulfate and alkylbenzene sulfonates include, but are not limited to, potassium hydroxide, potassium chloride, calcium chloride, calcium carbonate, and barium chloride. Suitable flocculants include, but are not limited to, potassium aluminum sulfate; sodium aluminum sulfate; aluminum ammonium sulfate; calcium chloride; sodium phosphate; aluminum hydroxide; aluminum chloride; ferric chloride; ferric sulfate; ferrous sulfate; sodium silicate; calcium silicate; calcium phosphate; zinc chloride; calcium carbonate; calcium bicarbonate; potassium sulfate; magnesium phosphate; acrylamide; acrylic acid; aluminum chlorohydrate; polyaluminum chloride; tannin; formaldehyde; melamine; N,N-dimethylaminoethyl acrylate methyl chloride; N,N-dimethylaminoethyl methacrylate methyl chloride quaternary salt; and polydiallyldimethylammonium chloride.

[0081] It should be understood herein that the cell disruptor undergoes a change in the precipitation process. For example, when the cell disruptor is sodium dodecyl sulfate (SDS), the precipitant may be potassium hydroxide (KOH), where the sodium cation is replaced by potassium as part of the precipitation process, and the resulting potassium dodecyl sulfate precipitates because it is insoluble in the reaction mixture. The dodecyl sulfate cation is functionally the cell disrupting portion of SDS and remains intact in this process.

[0082] In yet another embodiment of the above method, the cell disruptor may be one or more of sodium dodecyl sulfate (SDS) and sodium deoxycholate, and removal of the cell disruptor may include anionic adsorption. Anionic adsorption may include adding a suitable positively charged adsorbent in a suitable amount over a suitable time, followed by removal of the adsorbent. Anionic adsorption may further include flowing the reaction mixture through a column or filter packed with a suitable positively charged adsorbent at a suitable flow rate.

[0083] In yet another embodiment of the above method, the cell disruptor may be benzalkonium chloride, and removal of the cell disruptor includes cation adsorption. Cation adsorption may include adding a suitable negatively charged adsorbent in a suitable amount over a suitable time, followed by removal of the adsorbent. Cation adsorption may further include flowing the reaction mixture through a column or filter packed with a suitable negatively charged adsorbent at a suitable flow rate.

[0084] In yet another embodiment of the above method, the cell disruptor may be one or more of the Triton X100®, Triton X114®, Brij®, and Tween® surfactants, and removal of the cell disruptor includes micelle phase separation. Micelle phase separation may include changing the temperature of the reaction mixture so that the temperature of the reaction mixture exceeds the cloud point of the cell disruptor. Micelle phase separation may include centrifugation of the reaction mixture to obtain the desired phase separation.

[0085] In further embodiments of the method, the cell disruptor may be one or more of sodium dodecyl sulfate (SDS), benzalkonium chloride, Triton X100®, Triton X114®, Brij® surfactant, Tween® surfactant, sodium deoxycholate, and alkylbenzene sulfonates, and removal of the cell disruptor may include one or more of a combination of hydrophobic adsorption and dilution and tangential flow filtration (TFF). Hydrophobic adsorption may include adding a suitable hydrophobic adsorbent over a suitable amount of time, followed by removal of the adsorbent. Hydrophobic adsorption may further include flowing the reaction mixture at a suitable flow rate through a column or filter packed with a suitable hydrophobic adsorbent. Removal by dilution and TFF may include diluting the reaction mixture so that the cell disruptor can be removed by utilizing tangential flow filtration through a suitable molecular weight cut-off (MWCO) TFF filter, which allows the cell disruptor to pass through the retention solution containing Fukan, by reducing the cell disruptor to below its critical micelle concentration. Dilution and removal by TFF may include dialysfiltration of the reaction mixture through a TFF filter using an appropriate diavolute.

[0086] The method may further include adding a chelating agent to the reaction mixture to chelate free polyvalent cations in the reaction mixture. The chelating agent may be added after providing the starting Fukan composition but before removing the cell disruptor. The method may further include controlling the reaction of an oxidizing agent in the reaction mixture. Controlling the reaction of an oxidizing agent may include adding an oxidizing agent-reaction stopper to the reaction mixture before or after removing the cell disruptor.

[0087] The method described above may include adding a bacteriostatic agent to the reaction mixture. The bacteriostatic agent may be added after providing the starting Fukan composition but before removing the cell-disrupting agent. Suitable bacteriostatic agents include, but are not limited to, sodium sulfite, ethylenediaminetetraacetic acid (EDTA), benzalkonium chloride, ethanol, and thiourea.

[0088] Suitable chelating agents include, but are not limited to, ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin, and citric acid. Suitable oxidizing agents / reaction termination agents include, but are not limited to, sulfites, nitrites, and phosphates. As is clear from the above, some of the listed compounds may have more than one function in the above method.

[0089] Suitable hydrophobic adsorbents include, without limitation, activated carbon, diatomaceous earth, acrylic ester nonionic resins, polystyrene nonionic resins, and styrene-divinylbenzene (DVB) nonionic resins. Suitable anion adsorbents include, without limitation, styrene-DVB resins having amine functional groups, methacrylate resins having amine functional groups, methyl methacrylate resins having amine functional groups, butyl methacrylate resins having amine functional groups, agarose resins having amine functional groups, dextran resins having amine functional groups, ceramic resins having amine functional groups, silicates having amine functional groups, and lipid removers (LRAs).

[0090] In some embodiments, the starting fucoidan composition may be provided as a solution. Examples of fucoidans suitable for treatment by the above method include, but are not limited to, fucoidan. The starting fucoidan composition may have a fucoidan concentration in solution greater than 0.1% w / v and less than 30% w / v. The cell disruptor may have a fucoidan concentration in solution greater than 0.1% w / v and less than 60% w / v. Impurities that can be removed by the above method include, but are not limited to, particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA.

[0091] liquid-liquid extraction Fukan in a starting fukan composition, such as a raw fukan composition containing undesirable levels of impurities, is subjected to liquid-liquid extraction. The method may include providing the starting fukan composition in an aqueous starting solution, mixing the aqueous starting solution with an organic solvent to obtain an aqueous-organic phase mixture having an aqueous portion containing purified / modified fukan and an organic portion containing hydrophobic impurities, separating the aqueous portion from the organic portion, and recovering the aqueous portion containing the purified / modified fukan.

[0092] The method may further include desalting the starting Fukan composition before mixing the aqueous starting solution with the organic solvent. Desalting may include dialyseptic filtration of the starting Fukan composition, which is an aqueous solution, through a molecular weight cutoff (MWCO) tangential flow filtration (TFF) filter. Dialysis filtration may include dialyseptic filtration of the starting Fukan composition with distilled water. The molecular weight cutoff TFF filter may have a molecular weight cutoff smaller than the molecular weight in the purified / modified Fukan or the desired separation point or target molecular weight of the Fukan, for example, molecular weight cutoffs of 5 kDa, 10 kDa, 30 kDa, 50 kDa, 70 kDa, 100 kDa, 200 kDa, 300 kDa, 500 kDa, or 1000 kDa. Dialysis filtration may further include pre-filtration of the starting Fukan composition through a suitable pre-filter to remove particulate matter.

[0093] Mixing the aqueous starting solution with an organic solvent may include shaking the aqueous organic solvent mixture, stirring the aqueous organic solvent mixture, exposing the aqueous organic solvent mixture to high shear, recirculating the aqueous phase to the organic phase, and recirculating the organic phase to the aqueous phase.

[0094] Separating the aqueous portion from the organic portion may include at least one of the following: centrifugation, decantation, separation funnel separation, and hydrodynamic flow separation.

[0095] Suitable organic solvents for use in this method include organic solvents with a relative polarity of less than 0.765, such as heptane, isobutyl acetate, anisole, isopropyl acetate, 1-butanol, butyl acetate, methyl isobutyl ketone, pentane, 1-pentanol, ethyl acetate, ethyl ether, and propyl acetate. The organic phase may contain impurities such as, but are not limited to, lipids, fatty acids, phlorotannins, proteins, fucoxanthin, and / or chlorophyll.

[0096] diafiltration The fukan in a starting fukan composition, such as a raw fukan composition containing undesirable levels of impurities, is subjected to diafiltration. The method may include subjecting the starting fukan composition in the starting solution to diafiltration through a first tangential flow filter using a chelating agent solution to produce a filtration product of a first retained fukan composition and chelated cation components, and subjecting the first retained fukan composition to diafiltration through a second tangential flow filter using a secondary diafiltration solution to separate the first retaining solution chelating agent from the first retained fukan composition to produce a second retained fukan composition containing the desired purified / modified fukan. Subjecting the first retained fukan composition to diafiltration through a second tangential flow filter may include subjecting the first retained fukan composition to diafiltration through a first flow filter. That is, the same filter can be used in both diafiltration processes.

[0097] Subjecting the starting Fukan composition to dialysis filtration may include pre-filtration of the starting Fukan composition through a pre-filter to remove undesirable particulate matter. Subjecting the starting Fukan composition to dialysis filtration with a chelating agent may include subjecting the starting Fukan composition to dialysis filtration with one of ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin, or citric acid.

[0098] The starting fukan composition may have a fukan concentration in solution greater than 0.1% w / v and less than 30% w / v. The chelating agent may have a concentration in solution greater than 0.1% w / v and less than 60% w / v. The resulting first and / or second retaining liquid composition may have a cation content consisting essentially of sodium and / or potassium.

[0099] Figure 1 shows a schematic diagram of the cation content modification system 1200 for obtaining a change in the cation content and / or the level of the starting fucoidan composition. The starting fucoidan composition in solution is supplied to the fucoidan container 1216 via the inlet supply line 1202. The fucoidan of the starting material in a suitable solvent can be pre-filtered through a pre-filter 1204 to remove undesirable particulate matter. The gauge of the pre-filter is typically greater than the largest polymer molecules separated by the cation content modification system 1200.

[0100] The TFF inflow pump 1214 pumps the starting Fukan composition to the TFF filter 1210 via the TFF supply line 1212. The TFF filter 1210 is typically designed so that the supplied inflow fluid passes through the filter on its retaining liquid side, with the filtered product exiting through one outlet line and the treated inflow being supplied as a cassette with the retaining liquid via another outlet line. The TFF inflow pump 1214 applies a certain level of pressure to the TFF filter 1210 between its retaining liquid side and the filtered product side. In Figure 1, the retaining liquid of the TFF filter 1210 is returned to the Fukan container 1216 via the TFF retaining liquid return line 1218 and the TFF retaining liquid valve 1217, while the filtered product generated via the TFF filtered product outlet line 1219 is either used outside the cation content modification system 1200 or discarded.

[0101] While the TFF inflow pump 1214 recirculates the pre-filtered fucoidan and retaining solution to the TFF filter 1210, a chelating agent, for example, but not limited to ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin, or citric acid, may be added to the starting fucoidan composition in the fucoidan container 1216 from the first diafiltration solution container 1220 via the first diafiltration solution supply line 1225. The chelating agent is used both to replenish the solvent lost via the filtration product on the TFF filtration product outflow line 1219 and / or to ensure that a predetermined diavolume number of inflow fucoidan and chelating agent circulates over the TFF filter 1210. The chelating agent sequesters cations, particularly polyvalent cations, in the starting fucoidan composition as chelates, which then pass through the TFF filter 1210 to become filtration products. The chelating agent can be added in a pulsed manner by controlling the first diafiltration solution valve 1224. In other embodiments, the chelating agent may be added in a continuous mode. The number of diavolutes of the chelating agent to be processed on the TFF filter 1210 may be predetermined. The process may be continued for a predetermined period, for example, about 1 to about 6 hours, about 3 to about 12 hours, or about 10 to about 24 hours. The process may be continued within a predetermined number of diavolutes of the chelating agent, for example, about 1 to about 4 diavolutes, about 3 to about 6 diavolutes, about 5 to about 10 diavolutes, or about 7 to about 20 diavolutes. The process may be continued and the cation content of the Fukan container 1216 may be measured. The TFF process may be terminated when a desirable cation content is achieved, for example, a cation content in which the included polyvalent cations are less than 10 ppm, less than 1 ppm, less than 0.1 ppm, or less than 0.01 ppm. Using the first diafiltration solution, a first retained Fukan composition with a modified cation content is obtained by diafiltration of the starting Fukan composition in the solution through the TFF filter 1210.

[0102] The next step in the above process is to remove any remaining chelating agent from the first retained Fukan composition in the Fukan container 1216. This can be done by closing the first diafiltration solution valve 1224 and the outflow valve 1206 of the cation content changing system, allowing the secondary diafiltration solution from the second diafiltration solution container 1230 to enter the Fukan container 1216 via the second diafiltration solution supply line 1235 and the second diafiltration solution valve 1234. Subsequently, the mixture in the Fukan container 1216 is subjected to TFF through the TFF filter 1210 as described above, via the TFF supply line 1212, the TFF inflow pump 1214, the TFF retaining solution return line 1218, and the TFF retaining solution valve 1217. The secondary diafiltration solution may, for example, include one or more of deionized water, a bacteriostatic agent solution, and a salt, but are not limited to these. The bacteriostatic agent may be, for example, sodium sulfite, EDTA, benzalkonium chloride, ethanol, or thiourea. Suitable salts include, but are not limited to, sodium chloride, potassium chloride, sodium phosphate, ammonium bicarbonate, and phosphate-buffered saline.

[0103] The secondary diafiltration solution is used both to replenish the solvent lost via the filtration product on the TFF filtration product outflow line 1219 and / or to ensure that a predetermined number of diavolutes of the first retaining Fukan composition and the secondary diafiltration solution circulate through the TFF filter 1210. The secondary diafiltration solution can be added in pulsed mode by controlling the second diafiltration solution valve 1234. In other embodiments, the secondary diafiltration solution may be added in continuous mode. The number of diavolutes of the secondary diafiltration solution to be processed by the TFF filter 1210 may be predetermined. The process may continue for a predetermined period, for example, about 1 to about 6 hours, about 3 to about 12 hours, or about 10 to about 24 hours. This process may be continued with a predetermined number of diavolutes of chelating agent, for example, within the range of about 1 to about 4 diavolutes, about 3 to about 6 diavolutes, about 5 to about 10 diavolutes, or about 7 to about 20 diavolutes, until the cation content is such that it contains polyvalent cations of, for example, less than 10 ppm, less than 1 ppm, less than 0.1 ppm, or less than 0.01 ppm. The process may be continued and the residual chelating agent concentration in the Fukan container 1216 may be measured, and the TFF process may be terminated when an appropriate low residual chelating agent concentration, for example, less than 10 ppm, less than 1 ppm, less than 0.1 ppm, or less than 0.01 ppm, is achieved. The second retained Fukan composition obtained in the Fukan container 1216 contains the purified / modified Fukan product obtained by processing with the cation content modification system 1200. If desired, the second retained Fukan composition obtained in the Fukan container 1216 may be removed from the Fukan container 1216 via the cation content modification system outflow line 1208.

[0104] supercritical fluid extraction Fucan in a fucan composition, such as a starting fucan composition containing undesirable levels of impurities, is subjected to supercritical fluid extraction. The method may include placing the starting fucan composition in a supercritical extraction apparatus, applying a suitable pressure of more than 70 bar to the starting fucan composition in the supercritical extraction apparatus, heating the starting fucan composition in the supercritical extraction apparatus to a suitable temperature of more than 30°C, filling the supercritical extraction apparatus with supercritical fluid to produce a supercritical fluid containing purified / modified fucan and extracted impurities, removing the supercritical fluid containing the extracted impurities after a predetermined time, and recovering the purified / modified fucan.

[0105] Filling the supercritical extraction apparatus with a supercritical fluid may include filling the supercritical extraction apparatus with carbon dioxide. 2% v / v to 10% v / v ethanol may be added to the supercritical carbon dioxide. In some embodiments, approximately 5% v / v ethanol may be added to the supercritical carbon dioxide as a cosolvent. Alternative supercritical fluids to carbon dioxide used in this method include, but are not limited to, ethanol, ethane, hydrochloric acid, hydrofluoric acid, sulfuric acid, and nitric acid.

[0106] Applying appropriate pressure to the starting fukan may include applying a pressure of about 70 bar to about 2000 bar to the starting fukan composition. Exposing the starting fukan composition to an appropriate temperature may include exposing the starting fukan composition to a temperature of about 30°C to about 300°C.

[0107] Removing the supercritical fluid containing extracted impurities after a predetermined time may include removing the supercritical fluid after approximately 5 minutes to approximately 50 hours, for example, approximately 10 minutes to approximately 1 hour, approximately 30 minutes to approximately 5 hours, approximately 1 hour to approximately 24 hours, or approximately 5 hours to approximately 48 hours.

[0108] The method may further include desalting the starting Fukan composition before placing it in a supercritical fluid extractor. Desalting may include dialyseptic filtration of the starting Fukan composition, which is an aqueous solution, through a molecular weight cutoff (MWCO) tangential flow filtration (TFF) filter. Dialysis filtration may include dialyseptic filtration of the starting Fukan composition with distilled water. The molecular weight cutoff TFF filter may have a molecular weight cutoff smaller than the molecular weight in the purified / modified Fukan or the desired separation point or target molecular weight of the Fukan, for example, a molecular weight cutoff of 50 kDa, 70 kDa, 100 kDa, 200 kDa, 300 kDa, 500 kDa, or 1000 kDa. Dialysis filtration may further include pre-filtration of the starting Fukan composition through a suitable pre-filter to remove particulate matter.

[0109] Chemical structure modification The methods, systems, etc., described herein may include chemical structural modification of fukan, particularly fukan in fukan compositions. Chemical structural modification may include the removal of functional groups derived from fukan, such as O-acetyl, N-acetyl, methoxy, hydroxy, carboxylic acid, and / or sulfuric acid functional groups derived from the fukan structure. Such chemical structural modification may include the use of a wide range of chemical reagents, such as acids, bases, surfactants, and / or oxidizing agents.

[0110] Tangential flow filtration Some of the methods discussed herein utilize tangential flow filtration (TFF). As with typical identification of tangential flow filtration (TFF) filters, a given nominal molecular weight cutoff (MWCO) value of a TFF filter selectively retains on its retained side solutions containing molecules with molecular weights and / or sizes greater than those that do not cross the filter barrier and, therefore generally, those that cross / penetrate the barrier to the permeate side. Thus, the molecular weight cutoff value of a TFF filter is typically not absolute for any given polymer or nominal fractionation value. A given TFF filter will pass through or retain some molecules both above and below its nominal molecular weight cutoff value. The actual selectively selected values ​​and effects of a nominal TFF filter for a particular polymer can be routinely determined for that particular polymer.

[0111] Many factors can influence the permeation behavior of the TFF filter. These factors may depend on the TFF filter itself or on the attributes of the target polymer. For example, the folding behavior and folding structure of the target polymer can affect whether or not it crosses the MWCO barrier of the TFF filter. Regarding the TFF filter itself, as is known, many factors can influence its permeation behavior. For example, the manufacturing method can produce a variety of pore sizes within a particular TFF filter, and such variety may include both pores larger and smaller than the nominal MWCO. Therefore, a TFF filter with a nominal fractional molecular weight value will substantially allow / retain molecules at that nominal fractional molecular weight value, but will also allow / retain some molecules below and / or above that value.

[0112] Gel permeation chromatography Gel permeation chromatography was used to evaluate the molecular weight distribution obtained for the examples. Numerous different parameters, columns, and standards are available for use in gel permeation chromatography, resulting in a diverse range of instrumentation configurations for molecular weight analysis. For molecular weight measurements in this specification, GPC was performed using the following parameters: The mobile phase was a 0.1 M sodium nitrate stream at 0.6 mL / min. The column compartment and detector were at 30°C. A Waters 2414 differential refractive index detector was used for detection.

[0113] Suitable GPC columns include GPC columns compatible with aqueous solvents, such as columns packed with at least one of the following: sulfonated styrenedivinylbenzene, acrylate copolymer networks having NH functional groups, modified silica, and hydroxylated polymethacrylate gels. For the analysis described herein, three columns were used in series: one guard column with an inner diameter (ID) of 6 mm and a length of 40 mm packed with a hydroxylated polymethacrylate gel with a particle size of 6 μm; a first 300 mm analytical GPC column with an ID of 7.8 mm packed with a hydroxylated polymethacrylate gel with a particle size of 12 μm and an effective molecular weight range of approximately 50 kDa to 5,000 kDa; and a second 300 mm analytical GPC column with an ID of 7.8 mm packed with a hydroxylated polymethacrylate gel with a particle size of 10 μm and an effective molecular weight range of approximately 1 kDa to 6,000 kDa. The total effective molecular weight range of the column configuration was approximately 1 kDa to 6,000 kDa. An example of this column configuration could be an Ultrahydrogel® Guard column, an Ultrahydrogel® 2000 column, and an Ultrahydrogel® Linear column connected in series.

[0114] Traceable standards from American Polymer Standards Corporation, namely DXT3755K (peak molecular weight = 2164 kDa), DXT820K (peak molecular weight = 745 kDa), DXT760K (peak molecular weight = 621 kDa), DXT670K (peak molecular weight = 401 kDa), DXT530K (peak molecular weight = 490 kDa), DXT500K (peak molecular weight = 390 kDa), DXT270K ( Sample flow was quantified against a standard curve including Dextran 3755kDa (peak molecular weight = 196kDa), DXT225K (peak molecular weight = 213kDa), DXT150K (peak molecular weight = 124kDa), DXT55K (peak molecular weight = 50kDa), DXT50K (peak molecular weight = 44kDa), and DXT5K (peak molecular weight = 4kDa) (the peak molecular weights of these standards range from approximately 4kDa to approximately 2,200kDa). The standard curve used may include, for example, at least one of Dextran 3755kDa, Dextran 50kDa, and Dextran 55kDa, as well as 3 to 6 additional traceable standards discussed herein, with the calibration point being the peak molecular weight of the calibration sample used. Exemplary calibration curves may consist of DXT3755K, DXT820K, DXT530K, DXT500K, DXT225K, and DXT55K. The columns used herein had a total effective molecular weight range that encompassed and extended beyond the peak molecular weight range of the standard used for Fukan quantification.

[0115] The molecular weights described herein for fucoidan polymers are molecular weight values ​​in which there is always a distribution of higher and lower molecular weight molecules, where the amount or proportion increases or decreases as the molecular weight increases or decreases away from a particular molecular weight. The distribution may, but is not required to, have a general Gaussian shape or a distorted Gaussian shape.

[0116] The results in the tables herein include abbreviations used for specific characteristics of molecular weight distribution. Gel permeation chromatography is denoted as GPC, peak retention time as PRT, peak molecular weight as PMW, weight-average molecular weight as WAMW, number-average molecular weight as NAMW, percentage distribution as %dist, molecular weight as MW, polydispersity index as PDI, and fractionation molecular weight as MWCO.

[0117] Diseases and Conditions fibrous adhesions Fibrous adhesions are a type of scarring that forms between two parts of the body, usually after surgery (surgical adhesions). Fibrous adhesions can cause serious problems. For example, fibrous adhesions involving the female reproductive organs (ovaries, fallopian tubes) can cause infertility, painful intercourse, and severe pelvic pain. Fibrous adhesions that occur in the intestines can cause intestinal obstruction or blockage, and fibrous adhesions can also form in other places such as the heart, spine, and hands. In addition to surgery, fibrous adhesions can also be caused by conditions such as endometriosis, infection, chemotherapy, radiation, trauma, and cancer.

[0118] This specification discusses various types of fibrous adhesions. Terms such as surgical adhesion, post-surgical adhesion, postoperative adhesions, adhesions due to pelvic inflammatory disease, adhesions due to mechanical injury, adhesions due to radiation, adhesions due to radiation therapy, adhesions due to trauma, and adhesions due to the presence of foreign bodies all refer to the mutual adhesion of tissues through similar mechanisms and are all encompassed under the term fibrous adhesion.

[0119] Fibrous adhesions are a complex process in which tissues that are normally separate within the body erode and grow into each other. Surgical adhesions (also known as postoperative adhesions) develop from the tissue's otherwise normal wound healing response to trauma and are reported to occur in over two-thirds of all abdominal surgery patients (Ellis, H., Surg. Gynecol. Obstet. 133:497 (1971)). The outcomes of these fibrous adhesions are diverse and vary depending on other factors such as the surgical site and the site of the disease. Problems may include chronic pain, bowel obstruction, and an increased risk of death after cardiac surgery (diZerega, GS, Prog. Clin. Biol. Res. 381:1-18 (1993); diZerega, GS, Fertil. Steril. 61:219-235 (1994); Dobell, AR, Jain, AK, Ann. Thorac. Surg. 37:273-278 (1984)). In women of reproductive age, it is estimated that approximately 20% of all infertility cases are caused by fibrous adhesions, including those of the uterus, fallopian tubes, or ovaries (Holtz, G., Fertil. Steril. 41:497-507 (1984); Weibel, MA and Magno, G. Am. J. Surg. 126:345-353 (1973)).

[0120] The process of fibrous adhesion formation initially involves the establishment of a fibrin framework and the repair of normal tissue. In the normal repair process, fibrin degradation occurs as the mesothelium is repaired. However, in fibrous adhesion formation, the fibrin matrix grows during the proliferation of fibroblasts within the tissue network, leading to angiogenesis, and organized fibrous adhesions are established within approximately 3-5 days (Buckman, RF, et al., J.Surg.Res.21:67-76(1976); Raferty, AT, J.Anat.129:659-664(1979)). The inflammatory process includes neutrophil activation in the traumatic tissue, fibrin deposition and joining of adjacent tissues, macrophage infiltration, fibroblast proliferation within the area, collagen deposition, angiogenesis, and the establishment of permanent fibrous adhesion tissue.

[0121] Various methods have been attempted to prevent surgical adhesions. These include pharmacological approaches aimed at influencing the biochemical and cellular processes associated with surgical trauma, as well as septal techniques to isolate affected tissues. For example, peritoneal lavage, the use of heparinized solutions and procoagulants, modifications to surgical techniques such as microsurgery or laparoscopy, removal of talc from surgical gloves, the use of finer sutures, and the use of physical barriers (films, gels, or solutions) to minimize adhesion (apposition) to the serosal surface have all been tried. Currently, prophylactic treatment includes the prevention of fibrin deposition, reduction of inflammation (steroidal and nonsteroidal anti-inflammatory drugs), and removal of fibrin deposits.

[0122] Interventional attempts to prevent postoperative adhesion formation have included hydroflotation techniques or the use of barrier devices. Hydroflotation involves intravenously infusing a large amount of polymer solution, such as dextran (Adhesion Study Group, Fertil. Steril. 40:612-619 (1983)) or carboxymethylcellulose (Elkins, TE, et al., Fertil. Steril. 41:926-928 (1984)), into the surgical site to keep the organs involved isolated. Synthetic barrier membranes made from oxidized regenerated cellulose (e.g., Interceed®) or polytetrafluoroethylene (Gore-tex surgical membranes), as well as fully reabsorbable membranes made from a combination of modified hyaluronic acid / carboxymethylcellulose (HA / CMC) (Seprafilm®), have also been used to reduce postoperative adhesion formation in both animals and humans (Burns, JW, et al., Eur. J. Surg. Suppl. 577:40-48 (1997); Burns, JW, et al., Fertil. Steril. 66:814-821 (1996); Becker, JM, et al., J. Am. Coll. Surg. 183:297-306 (1996)). The success of these HA / CMC membranes is likely due to their ability to separate tissues during the peritoneal wound repair process when fibrous adhesions are formed. The aforementioned membrane was observed to form a transparent, viscous coating on the damaged tissue over a period of 3 to 5 days after application. This repair period coincides with the postoperative adhesion period (Ellis, H., Br.J.Surg. 50:10-16 (1963)). Unfortunately, the success observed with these methods was limited.

[0123] Peritonitis involves inflammation of the peritoneum. Peritonitis can cause severe symptoms, such as abdominal pain, abdominal tenderness, and abdominal wall guarding. Peritonitis may be associated with spontaneous inflammation, anatomic inflammation, and / or inflammation related to peritoneal dialysis. Infection may also be involved in peritonitis, for example, through perforation of a tubular organ, rupture of the peritoneum, spontaneous bacterial infection, and systemic infection. Peritonitis may not be caused by infection, for example, through leakage of sterile body fluid into the peritoneum and sterile abdominal surgery. Various attempts have been made to prevent and / or treat peritonitis, such as general symptomatic treatments including intravenous hydration, antibiotics, and surgery. There is still an unmet need for compounds, compositions, methods, and similar products (including delivery approaches) to suppress or otherwise treat and / or prevent peritonitis, preferably more effectively and with fewer side effects.

[0124] The purified / modified Fukan discussed herein may be used to treat fibrous adhesions in patients and may be included as an ingredient in a purified / modified Fukan medical device, purified / modified Fukan combination or purified / modified Fukan pharmaceutical configured and composed for treating fibrous adhesions, or may be a purified / modified Fukan medical device, purified / modified Fukan combination or purified / modified Fukan pharmaceutical configured and composed for treating fibrous adhesions. For example, it may be a purified / modified Fukan medical composition or medical device containing the purified / modified Fukan described herein dissolved in a physiological saline solution at an amount of about 0.02 mg / mL to about 100 mg / mL, for example, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.5 mg / mL, 0.9 mg / mL, 1 mg / mL, 2.5 mg / mL, 5 mg / mL or 7.5 mg / mL. The physiological saline solution may be, for example, Ringer's lactate solution USP (LRS), physiological saline solution, or physiological dextran solution.

[0125] The purified / modified Fukan medical compositions or medical devices described herein may be liquid compositions or liquid medical devices and may contain pharmaceutically acceptable excipients, such as buffers, stabilizers, preservatives, and adjuvants. Such purified / modified Fukan medical compositions or medical devices may be used to treat preoperative, intraoperative, or postoperative fibrous adhesions by administering the Fukan medical compositions or medical devices described in the preceding paragraph in an amount of about 0.01 mL / kg (per kilogram of patient or target body weight) to about 10 mL / kg or 15 mL / kg. Examples of the dosage of the purified / modified Fukan medical composition or medical device to the patient's surgical site include approximately 0.03 mL / kg, 0.1 mL / kg, 0.2 mL / kg, 0.4 mL / kg, 0.5 mL / kg, 0.6 mL / kg, 1 mL / kg, 1.2 mL / kg, 2 mL / kg, 3 mL / kg, 4 mL / kg, 5 mL / kg, 8 mL / kg, 10 mL / kg, or 15 mL / kg. In further embodiments, such purified / modified Fukan medical composition or medical device may be used to treat fibrous adhesions at any selected target site, such as lesions, abrasions, injury sites, surgical sites, and post-surgical sites, by administering approximately 0.04 mg / kg, 0.1 mg / kg to approximately 25 mg / kg, or 50 mg / kg. Some examples of such dosages include, for example, the fucane described herein, which includes approximately 0.04 mg / kg, 0.075 mg / kg, 0.1 mg / kg, 0.2 mg / kg, 0.5 mg / kg, 1 mg / kg, 1.3 mg / kg, 2 mg / kg, 3 mg / kg, 4 mg / kg, 5 mg / kg, 7.5 mg / kg, 8 mg / kg, 10 mg / kg, 15 mg / kg, 20 mg / kg, 25 mg / kg, or 50 mg / kg to the surgical site of the patient, including, for example, the purified / modified fucane described herein.The administration may be achieved, for example, by infusing the liquid medical composition or medical device over the entire target area; directing the liquid medical composition or medical device to one or more specific locations within the target area; spraying the liquid medical composition or medical device over the entire target area or to one or more specific locations; or by spraying or otherwise delivering the liquid medical composition or medical device via a trocar, catheter, endoscope, or other minimally invasive device, through an applicator which may be a spray applicator, to one or more specific locations identified by the surgeon or other expert as particularly prone to or at concern for the development of fibrous adhesions. In another embodiment, the administration may be performed after the surgical wound has opened and before the surgical procedure, during the surgical procedure, or after the surgical procedure and before the surgical wound has closed. If desired, the liquid medical composition or medical device may also be administered after the completion of the surgery (e.g., by syringe and needle), and may also be administered to non-surgical target sites. The patient's surgical site may be at least one of the following: pelvic cavity, abdominal cavity, dorsal cavity, cranial cavity, spinal cavity, abdominal cavity, thoracic cavity, pleural cavity, pericardial cavity, skin, joint, or muscle. Administration of the purified / modified Fukan medical composition or medical device into the patient's surgical site may be achieved in less than approximately 15 minutes, less than approximately 10 minutes, less than approximately 8 minutes, less than approximately 6 minutes, less than approximately 5 minutes, less than approximately 4 minutes, less than approximately 3 minutes, less than approximately 2 minutes, less than approximately 1 minute, less than approximately 45 seconds, less than approximately 30 seconds, less than approximately 20 seconds, less than approximately 15 seconds, less than approximately 10 seconds, or less than approximately 5 seconds.

[0126] Examples of administration of purified / modified Fukan medical compositions or medical devices to the surgical site include, after opening the surgical wound, during the surgery, before closing the surgical wound and / or after closing the surgical wound, in the following surgical procedures: cesarean section, microvascular free flap reconstruction, full-thickness skin grafting, VY stretch flap, fascial rotation flap, arthroplasty, mastectomy, necrotizing bone removal, and cup formation surgery. Surgical procedures, bone cutting surgical procedures, osteoplastic surgical procedures, patellar resection surgical procedures, synovectomy surgical procedures, capsule resection surgical procedures, tendon or ligament repair surgical procedures, tenolysis surgical procedures, surgical tendonectomy, fasciotomy surgical procedures, meniscus repair surgical procedures, vertebral resection surgical procedures, ethmoid resection surgical procedures, Caldwell-Look surgical procedures, dacryocystorhinostomy surgical procedures, lysis nasal synechia surgical procedures Procedures include: thymectomy, pulmonary dissection, lung resection, thoracoplasty, bilobectomy, portal hypertension surgery, splenectomy, esophagectomy, peritonitis surgery, gastrectomy, jejunostomy, laparoscopic cholecystectomy, laparoscopic common bile duct examination, gastrointestinal anastomosis, bariatric surgery, bowel resection and anastomosis, segmental hepatectomy, lobectomy, pancreaticotomy, pancreaticoduodenectomy, tumor resection, laparoscopic nephrectomy, cystectomy, and abdominal or pelvic adhesion dissolution surgery. Examples of surgical procedures that may involve the administration of purified / modified Fukan medical compositions or medical devices at the surgical site of procedures such as hysterosalpingography, hysterosalpingography, tubal reconstruction, laparoscopic surgery for ectopic pregnancy, joint replacement surgery, fracture repair surgery, hysterectomy, gallbladder removal surgery, cardiac bypass surgery, angioplasty, atherosclerotic plaque removal surgery, breast biopsy surgery, carotid endarterectomy surgery, cataract surgery, coronary artery bypass surgery, endometrial curettage surgery, hernia repair surgery, lumbar spine surgery, partial colectomy surgery, prostatectomy surgery, and tonsillectomy surgery.

[0127] General Cancer Information Cancer is the second leading cause of death in the United States, accounting for over 20% of all deaths. Cancer is a proliferative disorder characterized by the uncontrolled division of certain cells, which can lead to the formation of one or more tumors. Many methods are used to treat cancer, including surgery, radiation therapy, chemotherapy, and combinations thereof. Surgery is a relatively common method used for some localized tumors, but the possibility of tumor recurrence after tumor resection remains significant.

[0128] Treatment of cancer and other proliferative disorders has been limited by the potential for damage to or toxicity to non-cancerous healthy tissue. In radiotherapy and surgical procedures, the procedure has generally been limited to the tumor site and performed proximal to the tumor site. However, patients undergoing surgical removal of cancerous tissue may face significant risks (for example, in the removal of prostate or brain tumors, there is a significant risk of irreparable damage to surrounding major tissues, for instance, due to the potentially low need for resection of non-tumor tissue). Furthermore, focused radiation therapy, which has been the first-line treatment for prostate cancer, carries similar risks. In chemotherapy for cancer, drugs have been administered systemically, resulting in systemic exposure. While these drugs are designed to be toxic to cancer cells, they are also (generally) toxic to non-cancerous cells, resulting in patients becoming very ill when undergoing drug treatment for cancer. Through experience, oncologists can administer these drugs at doses that some patients can tolerate. However, these doses are often unsuccessful in treating cancer.

[0129] One problem associated with all methods of treating cancer is local recurrence of the disease. For example, approximately 700,000 Americans are diagnosed with localized cancer each year (approximately 64% of all cancer patients), and about 500,000 are treated surgically. Unfortunately, 32% of patients treated surgically experience recurrence after initial treatment (approximately 21% recur at the initial surgical site and 11% recur at a distant metastasis site). Approximately 100,000 patients die each year from localized cancer recurrence. This was particularly true for breast cancer, where 39% of patients undergoing breast tumor removal experienced local recurrence of the disease.

[0130] Staging is a method for determining the progression of cancer (solid tumors) in a patient. Using a simplified approach, patients are placed into one of three groups or stages based on the extent of their cancer progression.

[0131] Stage 1: The cancer can be treated by surgically removing part of the organ. This is also known as the resectable stage.

[0132] Stage 2: The cancer has progressed past the point where it can be surgically removed, but it remains confined to the organ itself.

[0133] Stage 3: The tumor has spread to other organs.

[0134] Many cancers are treated with antiproliferative agents, including, for example, 5-fluorouracil (Efudex®), vinca alkaloids (e.g., vincristine (Oncovin®)), anthracyclines (e.g., doxorubicin (Adriamycin®)), cisplatin (Platinol-AQ®), gemcitabine hydrochloride (Gemzar®), methotrexate, and paclitaxel. Some examples of toxicity associated with the antiproliferative agents methotrexate and paclitaxel are discussed elsewhere in this specification. Methotrexate has been used to treat several cancers, including, for example, bladder cancer, breast cancer, cervical cancer, head and neck cancer, liver cancer, lung cancer, and testicular cancer. Paclitaxel has been used to treat several cancers, including, for example, ovarian cancer, breast cancer, and non-small cell lung cancer (Compendium of Pharmaceuticals and Specialties, 35th Edition, 2000).

[0135] Toxicity from 5-fluorouracil includes cardiovascular toxicity such as myocardial ischemia; central nervous system toxicity such as euphoria, acute cerebellar syndrome, and ataxia; dermatological toxicity such as alopecia and dermatitis; gastrointestinal toxicity such as nausea, vomiting, and oral or gastrointestinal ulcers; hematological toxicity such as leukopenia, thrombocytopenia, and anemia; hypersensitivity toxicity such as hypersensitivity and contact hypersensitivity; ocular toxicity such as increased lacrimation, photophobia, and conjunctivitis; and other toxicity such as fever. 5-fluorouracil has been used to treat many cancers, including, for example, breast cancer, colorectal cancer, gastric cancer, liver cancer, bladder cancer, head and neck cancer, non-small cell lung cancer, ovarian cancer, pancreatic cancer, and prostate cancer (Compendium of Pharmaceuticals and Specialties Thirty-fifth Edition, 2000).

[0136] Toxicity from vincristine includes central nervous system toxicity such as seizures and hallucinations in children; dermatological toxicity such as alopecia; extravascular toxicity such as vesicant formation; gastrointestinal toxicity such as nausea, vomiting, constipation, and stomatitis; hematological toxicity such as bone marrow suppression; neurotoxicity such as peripheral neuropathy and autonomic neuropathy; ocular toxicity such as diplopia, transient blindness, and optic nerve atrophy; nephro / metabolic toxicity such as urinary retention, hyperuricemia, and bladder relaxation; respiratory toxicity such as shortness of breath; and other toxicity such as fever in children. This antiproliferative agent has been used to treat several cancers, including Hodgkin's disease, small cell lung cancer, Wilms' tumor, and testicular cancer (Compendium of Pharmaceuticals and Specialties Thirty-fifth Edition, 2000).

[0137] Toxicity from doxorubicin includes cardiovascular toxicity such as electrocardiogram abnormalities and cardiomyopathy; dermatological toxicity such as alopecia and nail changes; extravascular migration disorder toxicity such as blistering; gastrointestinal toxicity such as nausea, vomiting, and stomatitis; urogenital toxicity such as red discoloration of urine; hematological toxicity such as bone marrow suppression; hypersensitivity toxicity such as hypersensitivity and skin rash; ocular toxicity such as conjunctivitis; reproductive toxicity such as infertility; and other toxicity such as hyperuricemia. This antiproliferative agent has been used to treat several cancers, including breast cancer, small cell lung cancer, and ovarian cancer (Compendium of Pharmaceuticals and Specialties Thirty-fifth Edition, 2000).

[0138] Cisplatin toxicity includes cardiovascular toxicity such as electrocardiogram changes; dermatological toxicity such as hyperpigmentation; extravasation toxicity such as irritation; gastrointestinal toxicity such as nausea and vomiting; hematological toxicity such as bone marrow suppression and hemolytic anemia; hypersensitivity toxicity such as hypersensitivity reactions; neuromuscular toxicity such as peripheral neuropathy and acute degenerative brain disease; ophthalmic toxicity such as retrobulbar optic neuritis; otological toxicity such as hearing loss and tinnitus; nephrotoxicity / metabolic toxicity such as toxic nephropathy and hypokalemia; and other toxicity such as infertility. This antiproliferative agent has been used to treat several cancers, including, for example, bladder cancer, small cell lung cancer, ovarian cancer, testicular cancer, brain cancer, breast cancer, cervical cancer, head and neck cancer, hepatoblastoma, and thyroid cancer (Compendium of Pharmaceuticals and Specialties Thirty-fifth Edition, 2000). Toxicity from gemcitabine hydrochloride includes, for example, hematological toxicity such as bone marrow suppression; gastrointestinal toxicity such as nausea, vomiting, and stomatitis; hepatotoxicity such as transient elevation of serum transaminases; nephrotoxicity such as proteinuria, hematuria, hemolytic uremic syndrome, and renal failure; dermatological toxicity such as rash and alopecia; edema toxicity such as edema and peripheral edema; and other toxicity such as fever. This antiproliferative agent has been used to treat pancreatic cancer and non-small cell lung cancer (Compendium of Pharmaceuticals and Specialties Thirty-fifth Edition, 2000).

[0139] This study includes the prevention or treatment of localized cancers or solid tumors that can be treated, such as localized cancers or solid tumors of the prostate, breast, pancreas, liver, kidney, genitourinary system, brain, gastrointestinal system, respiratory system, and head and neck. The compositions described herein may prevent or treat cancer, including metastases, by delivering effective concentrations of purified / modified fucan to the tumor and / or metastases by diffusion or further systemic transport, and by enabling controlled release of purified / modified fucan to sites somewhat distant from the target tumor. Some of these cancers are discussed further in the following paragraphs.

[0140] prostate cancer Prostate cancer is a malignant tumor that develops in the cells lining the prostate gland. In the United States, an estimated 200,000 people will develop prostate cancer this year, and more than 30,000 will die from the disease. The mortality rate for new cases of prostate cancer is approximately 15%. The cancer may remain within the prostate or it may spread to surrounding tissues or distant sites (most often lymph nodes and bone). Prostate cancer usually spreads silently, and symptoms only appear when it progresses beyond the prostate. In several studies, the five-year survival rate for patients with prostate cancer when diagnosed and treated in the early stages was 94%.

[0141] Prostate cancer is often considered a disease of men over 50. In fact, 80% of men with prostate cancer are over 60 years old. The probability of a man being diagnosed with prostate cancer in his lifetime is about 1 in 10, roughly the same as the probability of a woman developing breast cancer. The number of reported new cases has increased significantly in recent years as a result of improved testing that can detect the disease early, often long before symptoms appear. The probability of developing prostate cancer in any given year increases with age, but increases significantly after the age of 50.

[0142] Current treatment options for prostate cancer depend on the stage of disease progression, the patient's age, and overall health. Elderly patients with only early-stage cancer or those with other more serious conditions may be treated conservatively, while older patients with advanced cancer may receive more aggressive treatment. Prostate cancer has been treated with a variety of methods, including radiation therapy (external beam or brachytherapy), hormone regression or castration (surgical or chemical), antiproliferative agents, surgery, and elective treatment (i.e., "careful observation"). Treatment does not guarantee a cure, and some have significant side effects.

[0143] Early-stage prostate cancer (i.e., tumor confined to the prostate) may be managed with "careful observation." Surgery for prostate cancer has been recommended for patients who are otherwise in good overall health and whose tumor is confined to the prostate. The general treatment for localized prostate cancer in men under 70 years of age was radical prostatectomy (i.e., surgical removal of the prostate).

[0144] Patients with localized prostate cancer are generally treated with external beam radiation (EBR). This radiation kills cancer cells and shrinks the tumor. EBR accounts for less than 20% of localized prostate cancer treatments, and approximately 50% of these patients experience post-radiation recurrence of the disease. Coupled with the detection of early-stage prostate cancer and increasing patient demand, the use of close-range radiotherapy (i.e., local radiotherapy) is expected to increase. In 1995, only 2.5% of newly diagnosed patients were treated with close-range radiotherapy. Close-range radiotherapy involves implanting radioactive metal "seeds" into the prostate tumor.

[0145] Widespread treatments for prostate cancer include testicular removal or hormone therapy. Both are used to suppress or stop the production of testosterone, which has been causing cancer growth. Approximately 20% of all prostate cancer patients undergo hormone withdrawal therapy. Hormone therapy includes goserelin acetate (Zoladex®) or leuprolide acetate (Lupron®). 5-fluorouracil is an antiproliferative agent used to treat prostate cancer.

[0146] Breast cancer In the United States, breast cancer is the most common cancer among women, with approximately 180,000 new cases diagnosed each year (male breast cancer accounts for about 5% of all diagnosed breast cancers). Only lung cancer surpasses breast cancer as a cause of death in women, with breast cancer accounting for approximately 50,000 deaths annually. One in eight American women (or about 13%) will develop breast cancer in their lifetime. Over the past decade, the most reported breast cancers have been small, primary (independently occurring; not caused by metastasis) tumors. Approximately 70-80% of newly diagnosed patients showed early-stage disease (stage 1 or 2), and the vast majority did not have axillary lymph node involvement.

[0147] Most breast cancers are carcinomas (i.e., malignant tumors that grow from epithelial tissue). Sarcomas or tumors arising from connective tissue, bone, muscle, or fat account for less than 1% of breast cancers. Also, most breast cancers (about 75%) are tubular carcinomas that occur in the tissue lining the milk ducts. A much smaller number of cancers (about 7%) are found in the lobules of the breast and are called lobular carcinomas. Almost all other forms of breast cancer are caused by Paget's disease (cancer of the ulna and nipple) and inflammatory cancers.

[0148] Treatment for breast cancer is complex and depends on many factors. Two important factors are the type of tumor and the stage of progression. Tumor characteristics are particularly helpful in dividing individuals into two groups: (1) those with a low risk of cancer recurrence, and (2) those with a high risk of cancer recurrence. Certain prognostic factors determine whether a patient falls into one of these groups. These factors include tumor size; the presence of estrogen and progesterone (ER / PR) receptors; the cell proliferation cycle (whether tumor cells are actively dividing or in the "S phase"); the presence of a protein known as "her-2-neu protein"; tumor malignancy, i.e., an indicator of differentiation or change in tumor cells; and tumor ploidy, i.e., the number of sets of genetic material within tumor cells.

[0149] Treatment for primary breast cancer without significant lymph node involvement involves mammary tumor removal and radiotherapy. More significant lymph node involvement may warrant mastectomy and removal of auxiliary lymph nodes. At this stage, the likelihood of metastasis and local recurrence is high. Treatment for metastatic disease is symptomatic and includes radiotherapy and chemotherapy, which suppress the immune system, damage cells, and reduce white blood cell counts. Antiproliferative agents, including, for example, 5-fluorouracil, doxorubicin, methotrexate, and paclitaxel, are approved for use in breast cancer.

[0150] Pancreatic cancer The pancreas is an organ of the digestive system located near the stomach and small intestine. It has two main functions: the production of enzymes and hormones. Pancreatic cancer can occur either exocrine (i.e., enzyme-producing) or endocrine (i.e., hormone-producing) within the pancreas (e.g., classic pancreatic adenocarcinoma).

[0151] Exocrine pancreatic cancer is a very serious health problem. In the United States, approximately 28,000 people are diagnosed with pancreatic cancer each year, while roughly the same number die from the disease. Pancreatic cancer occurs equally in men and women. Due to the difficulty in diagnosis, the inherent aggressive nature of pancreatic cancer, and the limited systemic treatment options available, only about 4% of patients diagnosed with pancreatic adenocarcinoma survive for five years. Pancreatic cancer is the fifth leading cause of cancer death, after breast cancer, lung cancer, colon cancer, and prostate cancer.

[0152] The choice of treatment for pancreatic cancer largely depends on the stage of the tumor. Possible treatments include surgery, antiproliferative agents, radiation, and biological therapy. Surgery is usually reserved for stage 1 patients where the cancer is considered resectable. In some cases, the combination of treatments such as radiation and antiproliferative agents administered before or after surgery can increase the patient's survival rate. Pancreatic cancer considered unresectable (usually stage II and above) may be treated with antiproliferative agents used in clinical trials. Antiproliferative agents such as gemcitabine or 5-fluorouracil have some effect on pancreatic cancer, and gemcitabine has been used as a symptomatic agent. The toxicity of these antiproliferative agents is discussed elsewhere in this specification. Radiation therapy, when used in combination with chemotherapy, has some effect on pancreatic cancer. Symptoms can sometimes be suppressed by radiation therapy alone. This form of treatment is also used for stage II and above pancreatic cancer.

[0153] Bladder cancer In 1998, more than 54,000 new cases of bladder cancer were diagnosed in the United States, and it was estimated that approximately 15,000 deaths were attributable to the disease. Bladder cancer is the fourth most common cancer among American men and the ninth most common cancer among American women. It occurs three times more frequently in men than in women. Bladder cancer is a major cause of disease and death in older men. The risk of bladder cancer increases sharply with age (80% of cases occur in people over 50), and more than half of all deaths from bladder cancer occur after age 70. In white people over 65, the annual incidence of bladder cancer is approximately 2 cases per 1,000 people, in contrast to 0.1 cases per 1,000 people under 65. The probability of a person developing bladder cancer in their lifetime is higher than 3%, but the probability of dying from bladder cancer is low (<1%). Bladder cancer is rare in people younger than 40 years old.

[0154] Recent studies suggest that certain genes and hereditary metabolic capabilities may play a role in bladder cancer. Transitional cell carcinoma (TCC) is the most common form of bladder cancer. TCC usually arises as a superficial (surface) mass on the stalk base, a papillary (verrucous) mass, or an lateral (growing outward) mass. However, in some cases, TCC may be attached to a broad base, or it may appear ulcerated (within a depression). Papillary TCC often begins as a hyperplastic area that later dedifferentiates or loses the characteristics of individual cells. Only about 10%–30% of papillary TCC progress to invasive cancer. In contrast, non-papillary forms of TCC are more likely to become invasive. As described, such TCC may appear ulcerated or flat. Flat, non-papillary TCC composed of undifferentiated epithelium is classified as carcinoma in situ (CIS or TIS). CIS tissue contains large, prominent nucleoli (round bodies within cells; involved in protein synthesis) and cells lacking normal polarity.

[0155] The treatment of bladder cancer depends on many factors, the most important of which are the type and stage of the tumor. Common treatments include transurethral resection (TUR), electrosurgery, laser surgery, intravesical treatment, antiproliferative agents, surgical treatment, cystectomy, and radiotherapy. Examples of antiproliferative agents used to treat bladder cancer include, for example, 5-fluorouracil, cisplatin, and methotrexate. The toxicity of the delayed antiproliferative agents 5-fluorouracil, cisplatin, and methotrexate is discussed elsewhere in this specification.

[0156] brain cancer Brain tumors are often inoperable, and more than 80% of patients die within 12 months of diagnosis. Approximately 18,000 new cases of primary intracranial (brain) cancer are diagnosed annually in the United States. Brain tumors account for about 2 percent of all adult cancers. More than 50 percent of brain tumors are high-grade gliomas (i.e., glioblastoma pleomorphoni and anaplastic astrocytoma). Patients with these tumors often suffer from severe disabilities, including motor impairment, seizures, and visual impairment.

[0157] Tumors that develop within brain tissue are known as primary brain tumors. Primary brain tumors are classified according to the type of tissue in which they develop. The most common type of brain tumor is the glioma, which develops within the glial (supporting) tissue. Other types include astrocytoma, brainstem glioma, ependymoma, and oligodendroglioma.

[0158] Surgical removal of brain tumors is recommended for most types and locations, but should be as complete as possible within the constraints of preserving neurological function. An exception to this rule is deep-seated tumors such as pontine gliomas, which are diagnosed based on clinical evidence and treated without initial surgery in approximately 50% of cases. However, in many cases, diagnosis is made by biopsy. Stereotactic biopsy can be used for lesions that are difficult to reach and resect. Patients with brain tumors that are rarely curable or unresectable should be considered candidates for clinical trials evaluating radiosensitizers, hyperthermia, or intratissue near-brightness radiotherapy used in combination with external beam radiotherapy to improve local control of the tumor, or for studies evaluating new drugs and bio-response modifiers.

[0159] Radiation therapy plays a major role in the treatment of most tumor types and can increase cure rates or extend disease-free survival. Radiation therapy may also be useful in treating recurrence in patients initially treated with surgery alone. Chemotherapy may be used before, during, or after surgery and radiation therapy. Similarly, recurrent tumors are also treated with chemotherapy. Cisplatin is an example of an antiproliferative agent used in the treatment of brain cancer. Examples of toxicity associated with this antiproliferative agent are discussed elsewhere in this specification.

[0160] restenosis Restenosis is a form of chronic vascular injury that leads to thickening of the vessel wall and loss of blood flow supplied by the vessel to the tissue. This inflammatory condition can occur in response to vascular reconstructive procedures, including any manipulations that reduce vascular occlusion. Therefore, restenosis is a major limiting factor that restricts the effectiveness of these procedures.

[0161] This study includes, for example, the prevention or treatment of restenosis by administering a therapeutically effective amount of oligonucleotide therapeutic agent and anti-inflammatory agent into a blood vessel. Suitable compositions may include a polymer carrier that can be surgically implanted at the site of restenosis or potential restenosis, or they may be injected via a catheter as a polymer paste or polymer gel. Suitable compositions may also include purified / modified Fucan as discussed herein.

[0162] arthritis Rheumatoid arthritis (RA) is a debilitating chronic inflammatory disease characterized by pain, swelling, synovial cell proliferation (pannus formation), and destruction of joint tissues. In advanced stages, the disease often damages vital organs and can be fatal. The disease involves multiple elements, including complex cytokine interactions of the immune system (macrophages / monocytes, neutrophils, B cells, and T cells) and synovial cell dysfunction and proliferation. Early and aggressive treatment with disease-modifying antirheumatic drugs (DMARDs), such as methotrexate, is recommended and will be discussed elsewhere in this specification.

[0163] Crystal-induced arthritis has been characterized by crystal-induced activation of macrophages and neutrophils within the joints, followed by severe pain that lasts for several days. The disease progresses with shorter intervals between episodes and a greater likelihood of the patient remaining in a pathological state. This disease has generally been treated symptomatically with nonsteroidal anti-inflammatory drugs (NSAIDs) such as diclofenac sodium (Voltaren®). These anti-inflammatory drugs have toxicities including central nervous system toxicity such as dizziness and headache; dermatological toxicity such as rash and itching; gastrointestinal toxicity such as aggravated ulcerative colitis and Crohn's disease; urogenital toxicity such as acute renal failure and renal papillary necrosis; hematological toxicity such as agranulocytosis, leukopenia and thrombocytopenia; hepatotoxicity such as elevated hepatic transaminases and hepatitis; and other toxicities such as asthma and hypersensitivity.

[0164] This study includes, for example, the prevention or treatment of rheumatoid arthritis by administering to a patient a therapeutically effective amount of an oligonucleotide therapeutic agent and optionally an anti-inflammatory agent. Suitable compositions include a polymer carrier that can be injected into the joint as a controlled-release carrier of the anti-inflammatory agent, and microparticles (integrated within the polymer carrier) as a controlled-release carrier of the oligonucleotide therapeutic agent. Suitable compositions may include purified / modified Fucan as discussed herein. Such polymer carriers may take the form of polymer microspheres, pastes, or gels.

[0165] Inflammatory state The compositions described herein can, if desired, suppress or treat neutrophil-mediated inflammatory conditions, including, for example, administering a composition containing an oligonucleotide therapeutic agent and an anti-inflammatory agent to a patient. Examples of such conditions include crystal-induced arthritis, osteoarthritis, non-rheumatic inflammatory arthritis, mixed connective tissue disease, Sjögren's syndrome, ankylosing spondylitis, Behçet's syndrome, sarcoidosis, psoriasis, eczema, inflammatory bowel disease, chronic inflammatory lung disease, neuropathy, and multiple sclerosis. Some of these diseases are discussed further in the following paragraphs.

[0166] Chronic inflammatory skin diseases (including psoriasis and eczema) Psoriasis is a common chronic inflammatory skin disease characterized by itchy, burning, tingling, easily bleeding, raised, thickened, and scaly lesions. These diseases have proliferative and angiogenic components in the later stages of the disease, but patients often have an associated arthritis condition. Symptoms may be treated with steroidal anti-inflammatory agents such as prednisone or antiproliferative agents such as methotrexate, which are discussed elsewhere herein. The compositions herein may be used for the suppression, or otherwise treatment and / or prevention of chronic inflammatory skin diseases, such as psoriasis and / or eczema.

[0167] The following provides some further representative examples of inflammatory diseases, including inflammatory arthritis, which can be treated with the compositions discussed herein, for example, arterial embolization in arteriovenous malformations (vascular malformations); menorrhagia; acute hemorrhage; central nervous system disorders; hypersplenism; inflammatory skin diseases such as psoriasis; eczematous diseases (atopic dermatitis, contact dermatitis, eczema); immunobullous diseases; and various conditions including rheumatoid arthritis, mixed connective tissue disease, Sjögren's syndrome, ankylosing spondylitis, Behçet's syndrome, sarcoidosis, crystal-induced arthritis, and osteoarthritis (all of which are characterized by inflamed, painful joints as a prominent symptom).

[0168] ischemia Ischemia or ischemia involves limitations in the blood supply, such as a lack of oxygen, glucose, and other components necessary for proper tissue function, leading to tissue damage and / or dysfunction. Ischemia can cause serious problems. For example, tissues may become oxygen-deprived and necrotic, and blood coagulation may occur. Various attempts have been made to prevent and / or treat ischemia. For example, blood flow restoration therapy or reperfusion therapy. However, blood flow restoration therapy involves the reintroduction of oxygen, which can cause further damage due to the generation of free radicals, potentially leading to reperfusion injury. Reperfusion injury can cause serious problems. The compositions described herein may be used to suppress ischemia and / or reperfusion injury, or otherwise to treat and / or prevent it.

[0169] Endotoxemia Endotoxemia is the presence of endotoxins in the blood. Endotoxemia can cause serious problems. For example, endotoxemia can lead to septic shock. The compositions described herein may be used to suppress or, otherwise, treat and / or prevent endotoxemia.

[0170] keloid scars Keloid formation is the healing of wounds with raised scars. These raised scars include abnormal fibrous scarring. Keloid formation can cause severe problems, such as pain and disfigurement. The compositions described herein may be used to suppress, or otherwise treat and / or prevent, keloid formation and the resulting raised scars.

[0171] Keloids (keloid scars) are a type of scar that proliferates and spreads over normal skin. Keloids involve abnormal collagen proliferation, including abnormal proliferation of type I and type III collagen. Keloids can cause severe problems, such as pain and itching, and can ulcerate if infected. Attempts have been made to treat or prevent keloids, including the use of surgery, bandaging, steroid injections, and laser therapy. The compositions described herein may be used to suppress keloids or, otherwise, to treat and / or prevent them.

[0172] dermatitis Dermatitis includes inflammation of the skin, including atopic dermatitis and contact dermatitis. For example, contact dermatitis involves localized rashes and / or skin irritation that develop after contact with foreign substances on the skin. For example, atopic dermatitis is a chronic, recurrent, pruritic skin disease. Atopic dermatitis is sometimes referred to as Benier's prurigo, neurodermatitis, endogenous eczema, flexural eczema, infantile dermatitis, childhood eczema, and acute simple prurigo (prurigo diathsique). Eczema is a form of dermatitis. Other types of dermatitis include cavernous dermatitis, seborrheic dermatitis (dandruff), dyshidrotic dermatitis (pompholyx), urticaria, vesicular dermatitis (bullous dermatitis), and popular urticaria. Dermatitis can cause serious problems. Examples include dry skin, skin rash, skin edema, skin redness, skin itching, crusting, cracking, blistering, exudation, and hemorrhage. Attempts have been made to treat or prevent dermatitis, including the use of corticosteroids and coal tar. The compositions described herein may be used to suppress, or otherwise treat and / or prevent, dermatitis, including atopic dermatitis, eczema, contact dermatitis, cavernous dermatitis, seborrheic dermatitis, dyshidrotic dermatitis, urticaria, vesicular dermatitis, and papular urticaria.

[0173] Rosacea Rosacea is a chronic disease or condition typically characterized by facial erythema. Rosacea can cause serious problems. For example, rosacea typically presents as redness of the forehead, nose, or cheeks, and may also cause redness of the neck, ears, scalp, and chest. For example, rosacea can cause further symptoms including telangiectasia, papules, pustules, and itchiness, and in advanced cases, rhinophyma (a red, lobulated nose) may develop. Subtypes of rosacea include erythematous telangiectasia, papular-pustular rosacea, phymatous rosacea, and ocular rosacea. Attempts have been made to treat or prevent rosacea, including the use of anti-inflammatory and antibiotic agents. The compositions described herein may be used to suppress, or otherwise treat and / or prevent, rosacea, including the subtypes of erythematous telangiectasia, papular-pustular rosacea, and ocular rosacea.

[0174] Medical devices, medical materials, combinations and pharmaceutical products The considerations herein also provide medical devices, combinations, and pharmaceutical products that include the compositions considered herein in a medical device, combination product, or pharmaceutically acceptable container. Such products may further include precautions attached to the container. These precautions are typically in the form prescribed by the competent authority regulating the manufacture, use, or sale of medical devices, medical materials, combinations, and pharmaceuticals or biopharmaceuticals, and thus the precautions reflect the authority's approval of the composition, for example, that purified / modified Fucan is approved as an antiproliferative or anti-inflammatory agent for administration to humans or animals for the treatment of proliferative or inflammatory diseases (e.g., inflammatory arthritis, restenosis, surgical adhesions, psoriasis, and peritonitis). Instructions for the use of purified / modified Fucan described herein may also be included. Such instructions may include information regarding patient administration and method of administration.

[0175] This application further relates to methods for producing the various elements considered herein, such as the purified / modified Fukan and the composition itself, as well as methods for using them, for example, for treating the conditions, diseases, etc. described herein.

[0176] This application includes medical devices, medical materials, pharmaceutical combination products and pharmaceutical products comprising purified / modified fucoidan and fucoidan compositions as described herein for treating fibrous adhesions, arthritis, psoriasis or other desired diseases. Such materials, etc., can be used in pharmaceuticals for treating fibrous adhesions such as surgical adhesions, arthritis, psoriasis or other desired diseases. Also provided are methods for producing and using such pharmaceuticals capable of alleviating symptoms associated with at least one of fibrous adhesions, arthritis, and psoriasis in patients, including human patients, by using a pharmaceutically effective amount of fucoidan, such as fucoidan as considered herein, in combination with pharmaceutically acceptable excipients or buffers. [Examples]

[0177] The following examples provide illustrative considerations of certain embodiments described herein, but the disclosure and claims are not limited thereto.

[0178] Example 1: Chemical structural modification An exudate-extract was obtained from Laminaria hyperborea. The exudate-extract was filtered, and small molecules were removed by (over) tangential flow filtration (TFF) using a 100 kDa filter. To obtain otherwise unmodified sample A, the obtained retainate was freeze-dried. The obtained retainate was diluted to 0.25 M NaOH by adding a 10 M NaOH solution and allowed to stand at room temperature for 16 hours. The obtained sample was then centrifuged using a 50 kDa filter, the obtained retainate was recovered and freeze-dried to obtain base-treated sample B. Proton magnetic resonance spectroscopy ( 1 Both the unmodified sample A and the base-treated sample B were analyzed by 1H-NMR. 1 The 1H-NMR spectrum is shown in Figure 2A.

[0179] Figure 2A shows the chemical structural modification of Fukan, but the broad peak with a chemical shift of approximately 2.0 ppm present in the unmodified sample A is not present in the base-treated sample B.

[0180] Furthermore, 2D 1 H- 13 Unmodified sample A and base-treated / modified sample B were analyzed by 14C heteronuclear multiple quantum coherence (HMQC). The HMQC spectra shown in Figure 2B were acquired at 70°C using a 600 MHz spectrometer equipped with a 5 mm cold probe, while suppressing solvent signals. Many scans of the HMQC spectra were acquired in the range of 10 to 30 ppm in the carbon dimension, with the number of scans increasing by 8 each time (256 to 512 scans per scan), and these scans were combined to produce the spectra shown in Figure 2B.

[0181] The HMQC spectrum for unmodified sample A has a cross peak corresponding to the O-acetyl group, indicated by the circled signal in Figure 2B. This cross peak is absent in the spectrum for base-treated sample B. This indicates that the acetyl group was removed from Fucan, and therefore, that the NaOH treatment chemically altered the structure of Fucan in base-treated sample B.

[0182] Example 2 Physically Induced Aggregation A starting solution was obtained by dissolving brown powdered fucoidan raw material in distilled water at approximately 10% w / v. Sodium chloride was added to the starting solution to produce a mixture with a final sodium chloride concentration of approximately 0.1 M. The mixture was heated to near boiling point for 10-15 minutes. Treatment of the mixture at this temperature caused aggregation of suspended impurities and particulate non-fucoidan material. The mixture was centrifuged at 2300 G (centrifugal force) for 40 minutes to separate the fucoidan-containing solution from the aggregated non-fucoidan components. Visual inspection of the fucoidan-containing solution revealed a visual decrease in particulate matter and color. The freeze-dried portion of the fucoidan-containing solution contained an off-white powder with significantly less color than the raw material fucoidan used. For example, the loss of color can be quantified and compared by obtaining the ultraviolet / visible spectra of raw fucoidan at 10 mg / mL in water and purified / modified fucane at 10 mg / mL in water, determining the total absorbance in the visible region of the spectrum in the range of approximately 400 nm to approximately 700 nm, and confirming that the total absorbance of purified / modified fucane is reduced by at least approximately 5%, 10%, or 20% compared to the raw fucoidan.

[0183] Example 3: Solid-phase extraction The brown powdered fucoidan was added to a mixture of 0.5 M sodium hydroxide (NaOH) in 70% v / v ethanol / water at 40°C. The resulting reaction mixture was stirred and maintained at 40°C for 2 hours. Next, the reaction mixture was centrifuged to separate the solid purified / modified fucoidan from the 0.5 M sodium hydroxide (NaOH) in the supernatant of 70% v / v ethanol / water containing extracted impurities.

[0184] Visualization revealed that the purified / modified solid fucoidan contained significantly less color to the naked eye than the raw fucoidan. This loss of color indicates the removal of impurities such as phlorotannins, and since fucoidan does not contain chromophores, it becomes colorless when completely purified. For example, the loss of color can be quantified and compared by obtaining the ultraviolet / visible (UV / Vis spectrum) of 10 mg / mL raw fucoidan in water and the ultraviolet / visible (UV / Vis spectrum) of 10 mg / mL purified / modified fucoidan in water, determining the total absorbance in the visible region of the spectrum in the range of approximately 400 nm to approximately 700 nm, and confirming that the total absorbance of purified / modified fucoidan is reduced by at least approximately 5%, 10%, or 20% compared to the raw fucoidan.

[0185] Example 4 Chemically induced precipitation The raw material fucoidan composition was dissolved in distilled water at 15% w / v to form a starting solution. Observation revealed that the starting solution contained suspended particulate matter. Calcium chloride was added to the starting solution to a concentration of 0.5 M to produce a reaction mixture. To simulate known impurities in natural fucoidan, sodium alginate was added at a concentration of 5% w / w alginate / fucoidan, and starch was added at a concentration of 5% w / w starch / fucoidan. In this case, starch was used as a mimic for laminarin. 10M sodium hydroxide (NaOH) was added dropwise to the reaction mixture to adjust the pH to 7-8. This was done to avoid the decomposition of fucoidan in the reaction mixture. To avoid acidification of the reaction mixture by the subsequent addition of phosphoric acid, a minimum amount of 10M sodium hydroxide (NaOH) was added again to the reaction mixture. The addition of phosphoric acid brought the reaction mixture to 0.5M phosphate. This initiated the aggregation of suspended particulate matter and precipitated impurities via the action of calcium phosphate formed by the reaction of calcium chloride and phosphoric acid. The reaction mixture was left at room temperature for 10 minutes to allow aggregation to continue. The reaction mixture was centrifuged at 17568G for 17 minutes to separate the desired purified fucoidan from the aggregated impurities in the supernatant. The supernatant was visually inspected to qualitatively evaluate the color and the removal of particulate matter. Aliquots of the supernatant were analyzed by ultraviolet / visible absorption in the 300-800 nm region to evaluate the removal of non-fucoidan components that scatter and / or absorb light in the ultraviolet / visible spectral region. The aliquots of the supernatant were also freeze-dried to obtain the fucan content. The aliquots of the supernatant were also hydrolyzed with 3M hydrochloric acid (HCl) at 90°C and analyzed by high-performance anion exchange chromatography with pulsed amperometric detection (HPAE-PAD) to detect total carbohydrates and evaluate the removal of laminarin and alginates. To evaluate the removal of alginate by assessing the removal of laminarin, monomeric mannuronic acid, and monomeric guluronic acid, impurities were quantified against a monomeric glucose standard.

[0186] The analytical results of the starting material, fucoidan, and the obtained purified / modified fucoidan are shown in Table 1 below.

[0187] [Table 1]

[0188] Example 5 Chemically induced precipitation The raw material fucoidan composition was dissolved in distilled water at 15% w / v to form a starting solution. Observation revealed that the starting solution contained suspended particulate matter. To simulate known impurities in natural fucoidan, sodium alginate was added at a concentration of 5% w / w alginate / fucoidan, and starch was added at a concentration of 5% w / w starch / fucoidan. In this case, starch was used as a mimic of laminarin. 10M sodium hydroxide (NaOH) was added dropwise to the reaction mixture to adjust the pH to 7-8. This was done to avoid the decomposition of fucoidan in the starting solution in the event that the subsequent addition of aluminum sulfate would make the starting solution acidic. The reaction mixture was prepared by adding 0.1M aluminum sulfate to the starting solution. This initiated the precipitation of impurities and the aggregation of impurities and suspended particulate matter by the simultaneously formed aluminum hydroxide. The reaction mixture was left at room temperature for 10 minutes to allow aggregation to continue. The reaction mixture was centrifuged at 17568G for 17 minutes to separate the desired purified fucoidan from agglutinated impurities in the supernatant. The supernatant was visually inspected to qualitatively evaluate the color and particle removal. Aliquots of the supernatant were analyzed by ultraviolet / visible absorption in the 300-800 nm region to evaluate the removal of non-fucoidan components that scatter and / or absorb light in the ultraviolet / visible spectral region. The aliquots of the supernatant were also freeze-dried to obtain the fucoidan content. The aliquots of the supernatant were also hydrolyzed with 3M hydrochloric acid (HCl) at 90°C and analyzed by high-performance anion exchange chromatography (HPAE-PAD) with pulsed amperometric detection to detect total carbohydrates and evaluate the removal of laminarin and alginates. To evaluate the removal of alginates by evaluating the removal of laminarin, monomeric mannuronic acid, and monomeric guluronic acid, impurities were quantified against monomeric glucose standards.

[0189] The analytical results of the starting material fucoidan and the obtained purified / modified fucoidan are shown in Table 2 below.

[0190] [Table 2]

[0191] Example 6 Chemically induced precipitation The starting fucoidan composition was dissolved in distilled water at 15% w / v to form a starting solution. Observation revealed that the starting solution contained suspended particulate matter. To simulate known impurities in natural fucoidan, sodium alginate was added at a concentration of 5% w / w alginate / fucoidan, and starch was added at a concentration of 5% w / w starch / fucoidan. In this case, starch was used as a mimic for laminarin. Calcium chloride was added to the starting solution to a concentration of 0.5 M to produce a reaction mixture. This initiated the precipitation of alginate. 10 M sodium hydroxide (NaOH) was added dropwise to the reaction mixture to adjust the pH to 7-8. This was done to avoid the decomposition of fucoidan in the reaction mixture. The aluminum sulfate concentration of the reaction mixture was adjusted to 0.5 M. This initiated the aggregation of suspended particulate matter, calcium alginate precipitate, and other impurities via the action of calcium sulfate formed by the reaction of calcium chloride and aluminum sulfate, and also via the action of aluminum hydroxide formed from aluminum sulfate in the reaction mixture. The reaction mixture was left at room temperature for 10 minutes to allow aggregation to continue. The reaction mixture was centrifuged at 17568G for 17 minutes to separate the desired purified fucoidan from the aggregated impurities in the supernatant. The supernatant was visually inspected to qualitatively evaluate the color and removal of particulate matter. Aliquots of the supernatant were analyzed by ultraviolet / visible absorption in the 300-800 nm region to evaluate the removal of non-fucoidan components that scatter and / or absorb light in the ultraviolet / visible spectral region. The aliquots of the supernatant were also freeze-dried to obtain the fucoidan content. Aliquots of the supernatant solution were also hydrolyzed with 3M hydrochloric acid (HCl) at 90°C and analyzed by high-performance anion exchange chromatography (HPAE-PAD) with pulsed amperometric detection to detect total carbohydrates and evaluate the removal of laminarin and alginates. To evaluate the removal of alginates by evaluating the removal of laminarin, monomeric mannuronic acid, and monomeric guluronic acid, impurities were quantified against monomeric glucose standards.

[0192] The analytical results of the starting material fucoidan and the obtained purified / modified fucoidan are shown in Table 3 below.

[0193] [Table 3]

[0194] Example 7: Liquid-liquid extraction A starting fucoidan composition containing impurities is dissolved in distilled water to produce a 10 mg / mL aqueous starting solution. 20% v / v heptane is added to the aqueous starting solution containing the starting fucoidan composition, and the organic-aqueous mixture is then mixed under high shear for 30 minutes. After this mixing is completed, the organic-aqueous mixture is placed in a separation funnel to separate the organic phase from the aqueous phase. A more concentrated aqueous phase containing the desired fucoidan components settles at the bottom of the separation funnel, while a less concentrated organic phase containing impurities remains at the top of the separation funnel. The organic-aqueous mixture is allowed to stand in the separation funnel for 10 minutes. The aqueous phase is then decanted and recovered as the desired purified / modified fucoidan in solution. The fatty acid, phlorotannin, protein, fucoxanthin, and / or chlorophyll content of the purified / modified fucoidan in solution can be found to be about 30%, about 50%, or about 70% to about 100% less than that of the starting fucoidan composition.

[0195] Example 8: Liquid-liquid extraction A starting fucoidan composition containing impurities is dissolved in distilled water to produce a 10 mg / mL aqueous starting solution. 20% v / v 1-butanol is added to the aqueous starting solution containing the starting fucoidan composition, and the organic-aqueous mixture is then mixed under high shear for 30 minutes. After this mixing is completed, the organic-aqueous mixture is placed in a separation funnel to separate the organic phase from the aqueous phase. A more concentrated aqueous phase containing the desired fucoidan components settles at the bottom of the separation funnel, while a less concentrated organic phase containing impurities remains at the top of the separation funnel. The organic-aqueous mixture is allowed to stand in the separation funnel for 10 minutes. The aqueous phase is then decanted and recovered as the desired purified / modified fucoidan in solution. The fatty acid, phlorotannin, protein, fucoxanthin, and / or chlorophyll content of the purified / modified fucoidan in solution can be found to be about 30%, about 50%, or about 70% to about 100% less than that of the starting fucoidan composition.

[0196] Example 9: Liquid-liquid extraction A starting fucoidan composition containing impurities is dissolved in distilled water to produce a 10 mg / mL aqueous starting solution. 20% v / v ethyl acetate is added to the aqueous starting solution containing the starting fucoidan composition, and the organic-aqueous mixture is then mixed under high shear for 30 minutes. After this mixing is completed, the organic-aqueous mixture is placed in a separation funnel to separate the organic phase from the aqueous phase. A more concentrated aqueous phase containing the desired fucoidan components settles at the bottom of the separation funnel, while a less concentrated organic phase containing impurities remains at the top of the separation funnel. The organic-aqueous mixture is allowed to stand in the separation funnel for 10 minutes. The aqueous phase is then decanted and recovered as the desired purified / modified fucoidan in solution. The fatty acid, phlorotannin, protein, fucoxanthin, and / or chlorophyll content of the purified / modified fucoidan in solution can be found to be about 30%, about 50%, or about 70% to about 100% less than that of the starting fucoidan composition.

[0197] Example 10 Dialysis Filtration A fucoidan composition containing approximately 8% w / v of a starting solution or starting material was provided. The starting solution was filtered through a 0.22 micron filter. The cation content of the aliquots of the filtered starting solution was measured by inductively coupled plasma mass spectrometry (ICP-MS), and it was found to contain more than 0.01% w / w of aluminum / fucan, 10 -5 It was found to contain arsenic / fucan above %w / w and calcium / fucan above 0.01%w / w, all of which were undesirable levels for each cation. The starting solution was dialyzed through 4 divolumes of a solution of 0.1 M EDTA and 0.01 M sodium hydroxide (NaOH). Next, the obtained retained fucoidan solution was dialyzed through 2.5 divolumes of a solution of 5 mM sodium sulfite (Na2SO3) and 5 mM sodium chloride (NaCl). The obtained secondary retained fucoidan solution was assayed for cation content by ICP-MS. The results for the fucoidan composition of the starting material and the obtained purified / modified fucoidan are shown in Table 4 below.

[0198] [Table 4]

[0199] Example 11: Supercritical Fluid Extraction A fucoidan composition of approximately 100 g of solid starting material is provided. The solid is placed in a supercritical fluid extractor. The extractor is pressurized to 5800 psi, heated to 50°C, and then purged with supercritical carbon dioxide at a rate of 100 mL / min for 3 hours. The supercritical carbon dioxide is purged from the extractor, and the modified / purified solid fucoidan is recovered and analyzed for impurities. It can be found that the content of lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, free ions, bacteria, and / or DNA in the recovered modified / purified solid fucoidan is approximately 30% to approximately 100% less than that of the fucoidan composition of the starting material.

[0200] Example 12 Chemically induced precipitation, dissolution, and aggregation A starting material fucoidan composition, found to contain approximately 0.70% w / w total nitrogen, was dissolved in 15% w / v distilled water to form a starting solution. The presence of total nitrogen indicates the presence of undesirable impurities such as cellular components, DNA, proteins, and bacteria. Since nitrogen-containing impurities can chemically or ionically bond to fucoidan molecules, a decrease in total nitrogen in this case indicates that these nitrogen-containing impurities have been removed from the starting material fucoidan composition or fucoidan polymer.

[0201] Observation revealed that the starting solution contained suspended particulate matter. Calcium chloride was added to the starting solution to a concentration of 0.5 M to produce a reaction mixture. This initiated the precipitation of the solid portion, which was thought to contain impurities. Approximately 15 mL of 10 M sodium hydroxide (NaOH) was added dropwise to the reaction mixture to adjust the pH to 7-8. This was done to avoid the decomposition of fucoidan in the reaction mixture. The reaction mixture was adjusted to a phosphate concentration of 0.5 M (0.5 M phosphate) by adding phosphoric acid. This initiated the aggregation of suspended particulate matter and precipitated impurities via the action of calcium phosphate formed by the reaction of calcium chloride and phosphoric acid. The reaction mixture was centrifuged at 33,746 G for 5 minutes to separate the first purified / modified fucoidan from the aggregated impurities in the supernatant. The first purified / modified fucoidan was found to contain approximately 0.10% w / w total nitrogen. A portion of the first purified / modified fucoidan in the supernatant was further purified by dialysfiltration using a centrifugal filter with a MWCO of 100 kDa against 6 diavolutes of 5 mM sodium chloride (NaCl). The resulting first retained purified / modified fucoidan was found to contain approximately 0.08% w / w total nitrogen.

[0202] A further portion of the first purified / modified fucoidan was treated by adding a 1 M solution of sodium dodecyl sulfate as a cell disruptor to a concentration of 0.010 M. A 10 M solution of sodium hydroxide (NaOH) was added to a concentration of 0.26 M to make the mixture basic. The resulting reaction mixture was stirred at room temperature for about 30 minutes to obtain a turbid, light brown mixture.

[0203] After approximately 30 minutes, a 45% w / v potassium hydroxide (KOH) solution was added until the concentration reached approximately 0.04 M. Upon addition of potassium, undesirable impurities precipitated along with sodium dodecyl sulfate (SDS). A 48% w / v aluminum sulfate solution was then added until the concentration reached approximately 0.06 M. The formation of aluminum hydroxide caused undesirable impurities in the reaction mixture to aggregate. Sodium sulfite solid was added and dissolved to a concentration of 0.02 M to control the reaction of any oxidizing agents that may be present in the reaction mixture.

[0204] The resulting reaction mixture was stored in a refrigerator for approximately 16 hours, followed by centrifugation at 33,746 G for 5 minutes to separate the second purified / modified fucoidan from the aggregated impurities in the supernatant. The second purified / modified fucoidan was found to contain approximately 0.06% w / w total nitrogen. A portion of the second purified / modified fucoidan was further processed by dialysfiltration using a centrifugal filter with an MWCO of 100 kDa against 6 diavolutes of 5 mM sodium chloride (NaCl) to provide the second retained purified / modified fucoidan. The obtained second retained purified / modified fucoidan was found to contain approximately 0.03% w / w total nitrogen.

[0205] Example 13 Preparation of 5 types of purified / modified Fucan The methods discussed herein can be used, combined, modified, and reordered in any manner to obtain purified / modified fukan. Five types of purified / modified fukan were prepared using chemically derived precipitation and diafiltration combinations described in Examples 4, 5, 6, and 10, and their efficacy in medical and surgical applications was evaluated. These five types of fukan are referred to herein as Fukan 1 to Fukan 5. Fukan 1 and Fukan 2 were prepared in approximately 2 kg quantities using the methods described in Examples 4 and 10. Fukan 3 and Fukan 5 were prepared in approximately 30 g quantities using the methods described in Examples 4 and 10. Fukan 4 was prepared in approximately 1 kg quantities using the methods described in Examples 5 and 10. Fukan 1 to Fukan 5 were converted into solid purified / modified fukan by diafiltration into a low-conductivity salt solution followed by freeze-drying to obtain a white solid. Two further fucose compounds were extracted from brown algae and are referred to herein as fucose 6 and fucose 7. Fucan 6 was provided as a solid composition by FMCBioPolymer®. None of the above processes were used in the production of fucose 6. Fucan 7 was extracted from brown algae with near-boiling HCl. Some impurities were removed by selective precipitation using ethanol as a precipitating agent. After selective precipitation, the fucose was further precipitated with ethanol, centrifuged, and freeze-dried to recover the fucose as a solid composition. Fucan 7 was further treated in the manner discussed in Example 3, then dissolved in water, dialyzed against deionized water, and freeze-dried to obtain fucose 7 as a solid composition. The levels of fucose, galactose, sulfuric acid, and total counterions of fucose 1 to fucose 7 are calculated as discussed in Examples 14 and 15 below. These fucose compounds 1 to fucose 7 are discussed further below, for example, in Example 16 and Table 6.

[0206] Example 14 Measurement of corrected fucose content and corrected galactose content of Fucan 1 to Fucan 7 The solid Fukan composition was dissolved at 40 mg / mL in 72% w / w sulfuric acid and incubated in a 45°C water bath for 30 minutes. Next, the acid hydrolysate was diluted to a sulfuric acid concentration of 4% w / w (4% w / w sulfuric acid) in a high-pressure tube and incubated at 120°C for 60 minutes. The resulting second acid hydrolysate was diluted to a 1 / 333 concentration with distilled water and analyzed by high-performance anion exchange column chromatography with pulsed amperometric detection (HPAE-PAD). The analyte was separated by flowing a 10 mM sodium hydroxide (NaOH) eluent at 1.0 mL / min using an isocratic pump.

[0207] The uncorrected fucose content of Fucan was calculated by interpolation using a standard curve for fucose. The uncorrected galactose content of Fucan was calculated using the standard addition method. The corrected fucose content was calculated considering that one molecule of water is added during the hydrolysis of glycosidic bonds in fucose, and two hydroxyl groups are added during the hydrolysis of two sulfate ester bonds. The corrected galactose content was calculated considering that one molecule of water is added during the hydrolysis of glycosidic bonds.

[0208] The results of this analysis are shown in Table 5 below.

[0209] Example 15: Measurement of total sulfuric acid content, total counterion content, and total water content of Fucan 1 to Fucan 7 The solid Fukan composition was dissolved in deionized water and hydrolyzed under acidic conditions. The total sulfur and counterion content (%w / w) was analyzed by ICP-MS. The sulfur content was converted to sulfuric acid content by multiplying the sulfur content by the sulfuric acid-to-sulfuric acid molar ratio to obtain the sulfuric acid content (%w / w) of the purified / modified Fukan. The counterions observed in the purified / modified Fukan discussed here included potassium and sodium counterions. The results of this analysis are shown in Table 5 below.

[0210] The results (%w / w) of the total content of corrected fucose, corrected galactose, and sulfuric acid are also shown in Table 5 below. The total content of fucose, galactose, and sulfuric acid is calculated by adding together the values ​​of corrected fucose, corrected galactose, and sulfuric acid, and a more detailed and complete calculation, including the aspects described above, is shown in Equation 1 below. The total counterion content is calculated by adding together the total sodium and total potassium content.

[0211]

number

[0212] [Table 5]

[0213] Table 5 shows that purified / modified fukan containing impurities of less than approximately 12% w / w, less than approximately 10% w / w, less than approximately 9% w / w, less than approximately 8% w / w, less than approximately 7% w / w, less than approximately 6% w / w, less than approximately 5% w / w, less than approximately 4% w / w, less than approximately 3% w / w, or less than approximately 2% w / w can be prepared using the methods discussed herein.

[0214] Table 5 further shows that purified / modified fucane was produced with a total content of galactose, fucose, and sulfuric acid of approximately 77% w / w to approximately 87% w / w.

[0215] Table 5 further shows purified / modified fukan with a total counterion content of approximately 9% w / w to approximately 14% w / w relative to fukan.

[0216] The total water content of Fucan 1, Fucan 3, Fucan 4, and Fucan 5 was measured by loss on drying (LOD) at 104°C. The total water content was calculated to be 3.8% w / w, 2.4% w / w, 3.2% w / w, and 4.7% w / w for each purified / modified Fucan, respectively.

[0217] Example 16: Measurement of the molecular weight distribution of Fukan 3 and Fukan 4 Gel permeation chromatography was used to evaluate the molecular weight distribution obtained for the purified / modified Fucan 3 and Fucan 4. Numerous different parameters, columns, and standards are available for use in gel permeation chromatography, resulting in a variety of instrumentation configurations suitable for molecular weight analysis. For molecular weight measurements described herein, GPC was performed using the following parameters: The mobile phase was a 0.1 M sodium nitrate stream at 0.6 mL / min. The column compartment and detector were at 30°C. A Waters 2414 differential refractive index detector was used for detection.

[0218] Suitable GPC columns include GPC columns compatible with aqueous solvents, such as columns packed with at least one of the following: sulfonated styrenedivinylbenzene, acrylate copolymer networks having NH functional groups, modified silica, and hydroxylated polymethacrylate gels. For the analysis described herein, three columns were used in series: one guard column with an inner diameter (ID) of 6 mm and a length of 40 mm packed with a hydroxylated polymethacrylate gel with a particle size of 6 μm; a first 300 mm analytical GPC column with an ID of 7.8 mm packed with a hydroxylated polymethacrylate gel with a particle size of 12 μm and an effective molecular weight range of approximately 50 kDa to 5,000 kDa; and a second 300 mm analytical GPC column with an ID of 7.8 mm packed with a hydroxylated polymethacrylate gel with a particle size of 10 μm and an effective molecular weight range of approximately 1 kDa to 6,000 kDa. The total effective molecular weight range of the column configuration was approximately 1 kDa to 6,000 kDa. An example of this column configuration could be an Ultrahydrogel® Guard column, an Ultrahydrogel® 2000 column, and an Ultrahydrogel® Linear column connected in series.

[0219] Traceable standards from American Polymer Standards Corporation, namely DXT3755K (peak molecular weight = 2164 kDa), DXT820K (peak molecular weight = 745 kDa), DXT760K (peak molecular weight = 621 kDa), DXT670K (peak molecular weight = 401 kDa), DXT530K (peak molecular weight = 490 kDa), DXT500K (peak molecular weight = 390 kDa), DXT270K ( Sample flow was quantified against a standard curve including Dextran 3755kDa (peak molecular weight = 196kDa), DXT225K (peak molecular weight = 213kDa), DXT150K (peak molecular weight = 124kDa), DXT55K (peak molecular weight = 50kDa), DXT50K (peak molecular weight = 44kDa), and DXT5K (peak molecular weight = 4kDa) (the peak molecular weights of these standards range from approximately 4kDa to approximately 2,200kDa). The standard curve used may include, for example, at least one of Dextran 3755kDa, Dextran 50kDa, and Dextran 55kDa, as well as 3 to 6 additional traceable standards discussed herein, with the calibration point being the peak molecular weight of the calibration sample used. Exemplary calibration curves may consist of DXT3755K, DXT820K, DXT530K, DXT500K, DXT225K, and DXT55K. The columns used herein had a total effective molecular weight range that encompassed and extended beyond the peak molecular weight range of the standard used for Fukan quantification.

[0220] The results in Table 6 below include abbreviations used for specific characteristics of molecular weight distribution. Gel permeation chromatography is represented as GPC, peak molecular weight as PMW, weight-average molecular weight as WAMW, number-average molecular weight as NAMW, percentage distribution as %dist, and molecular weight as MW.

[0221] [Table 6]

[0222] Example 17 Preparation of high-purity Fukan composition by drying Approximately 100 mg of Fucan 1 was placed in a crucible. The crucible containing Fucan 1 was placed in a 105°C oven for 30 minutes to produce a further purified Fucan composition, hereafter referred to as Fucan 1'. The crucible containing the further purified Fucan 1' composition was removed from the oven and placed in a desiccator. The further purified Fucan 1' composition was analyzed for the total content of fucose and galactose by HPAE-PAD and for the total content of sulfur and counterions by ICP-MS under a moisture-free (absolutely dry) atmosphere. It was found that the total content of fucose, galactose, sulfuric acid, and counterions was 99.9% w / w or higher, meaning that impurities were less than 0.1%.

[0223] Example 18 Preparation of high-purity Fukan composition by drying Approximately 600 mg of Fucan 1 was placed on an aluminum dish of an Ohaus MB90 moisture analyzer. The analyzer was programmed to heat Fucan 1 at 105°C for 30 minutes to produce a further purified Fucan 1'' composition. The further purified Fucan 1'' composition was removed from the analyzer and placed in a desiccator. The sample was analyzed under a moisture-free (absolutely dry) atmosphere for the total content of fucose and galactose by HPAE-PAD and for the total content of sulfur and counterions by ICP-MS. It was found that the total content of fucose, galactose, sulfuric acid, and counterions was 99.9% w / w or higher, with impurities of less than 0.1%.

[0224] Example 19: Uterine horn fibrous adhesions treated with Fukan 1 To measure the efficacy of purified / modified Fucan 1 in suppressing surgical adhesions, the following double uterine horn (DUH) surgery was performed on both uterine horns of a total of two New Zealand White rabbits. Prior to surgery, the rabbits were weighed and then prepared for surgery with premedication of ketamine and xylazine.

[0225] A 0.33 mg / mL fucoidan solution was prepared in Ringer's lactate solution USP (LRS) and sterilized by filtration. All instruments were sterilized, and a sterile field was maintained during the surgery. The abdomen was cleaned, and access was made via a midline abdominal incision. The uterine horn was located, exposed outside the body, and injured by scraping. The abdominal wall near the scraped uterine horn was also scraped. A minimal amount of fucoidan solution was applied to the injured uterine horn and lateral wall region. The injured uterine horn and abdominal wall were positioned next to each other and fixed with sutures. Before closing the incision, approximately 15 mL / kg of fucoidan solution per rabbit's body weight was applied to the abdominal cavity. Adhesion was evaluated two weeks after the surgery. The length of uterine horn adhesions was measured with a ruler. The uterine horn adhesion coverage rate, which is the ratio of the length of adhesions to the total injured uterine horn length, was calculated. Equation 2: Adhesion coverage rate (%) = 100 × Uterine horn adhesion length ÷ Total damaged uterine horn length It was calculated as follows.

[0226] The same surgical method was applied to New Zealand White rabbits, but instead of fucoidan solution, approximately 15 mL / kg of the control, Ringer's lactate solution USP (LRS), was administered. The control group, administered LRS, had an adhesion coverage rate of 63%, calculated using Equation 2. Table 7 shows the results obtained using the methods discussed above for Fucan 2, a representative example of purified / modified Fucan. The results in the following tables are shown as a decrease in adhesion coverage rate compared to the control group.

[0227] Table 7 shows the results of treatment of six uterine horns by Fukan 1.

[0228] [Table 7]

[0229] As can be seen from the results in Table 7, the purified / modified Fukan described herein may be used to successfully treat postoperative uterine horn adhesions.

[0230] Example 20: Uterine horn fibrous adhesions treated with Fukan 4 To measure the efficacy of purified / modified Fucan 4 in suppressing surgical adhesions, the following double uterine horn (DUH) surgery was performed on both uterine horns of a total of four New Zealand White rabbits. Prior to surgery, the rabbits were weighed and then prepared for surgery with premedication of ketamine and xylazine.

[0231] A 3.75 mg / mL fucoidan solution was prepared in Ringer's lactate solution USP (LRS) and sterilized by filtration. All instruments were sterilized, and a sterile field was maintained throughout the surgery. The abdomen was cleaned, and access was made via a midline abdominal incision. The uterine horns were located, exposed, and scraped to induce damage. The abdominal wall near the scraped uterine horns was also scraped. 4 mL of fucoidan solution was applied directly to the damaged left uterine horn and lateral wall region, and 4 mL of fucoidan solution was applied directly to the damaged right uterine horn and lateral wall region. The damaged uterine horns and abdominal wall were positioned adjacent to each other and fixed with sutures. A drainage tube was placed in the abdominal cavity before closing the incision. The drainage tube was removed 48 hours postoperatively. Adhesions were evaluated two weeks after the surgery. The length of the uterine horn adhesions was measured with a ruler. The uterine horn adhesion coverage rate was calculated using Equation 2.

[0232] Three New Zealand White rabbits were subjected to the same surgical procedure, and instead of fucoidan solution, 4 mL / kg (4 mL per side) of Ringer's lactate solution USP (LRS) was administered as a control. The control group, which received LRS, had an adhesion coverage rate of 73%, as calculated using Equation 2. Table 8 shows the results obtained using the methods discussed above for four representative examples of purified / modified fucoidan compositions. The results in the following tables are shown as a decrease in adhesion coverage rate compared to the control group.

[0233] Table 8 shows the results of treatment of eight uterine horns by Fukan 4.

[0234] [Table 8]

[0235] As can be seen from the results in Table 8, purified / modified Fukan can be used to successfully treat postoperative uterine horn adhesions.

[0236] Example 21: Uterine horn fibrous adhesions treated with Fukan 6 To measure the efficacy of purified / modified Fucan 6 in suppressing surgical adhesions, the following double uterine horn (DUH) surgery was performed on both uterine horns of a total of four New Zealand White rabbits. Prior to surgery, the rabbits were weighed and then prepared for surgery with premedication of ketamine and xylazine.

[0237] A 0.33 mg / mL fucoidan solution was prepared in Ringer's lactate solution USP (LRS) and sterilized by filtration. All instruments were sterilized, and a sterile field was maintained during the surgery. The abdomen was cleaned, and access was made via a midline abdominal incision. The uterine horn was located, exposed outside the body, and injured by scraping. The abdominal wall near the scraped uterine horn was also scraped. A minimal amount of fucoidan solution was applied to the injured uterine horn and lateral wall region. The injured uterine horn and abdominal wall were positioned adjacent to each other and fixed with sutures. Before closing the incision, approximately 15 mL / kg of fucoidan solution per rabbit's body weight was applied to the abdominal cavity. Adhesion was evaluated two weeks after the surgery. The length of the uterine horn adhesion was measured with a ruler. The uterine horn adhesion coverage rate was calculated using Equation 2.

[0238] Four New Zealand White rabbits were subjected to the same surgical procedure, but instead of fucoidan solution, they were given approximately 15 mL / kg of the control, Ringer's lactate solution USP (LRS). The control group, administered LRS, had an adhesion coverage rate of 71%, calculated using Equation 2. Table 9 shows the results for Fukan 6 obtained using the method discussed above. The results in the following tables represent the decrease in adhesion coverage rate compared to the control group.

[0239] Table 9 shows the results of treatment of eight uterine horns by Fukan 6.

[0240] [Table 9]

[0241] As can be seen from the results in Table 9, Fucan 6, prepared using known methods and containing more than 50% non-Fucan components, was not effective in treating fibrous adhesions.

[0242] Example 22: Uterine horn fibrous adhesions treated with Fukan 7 To measure the efficacy of purified / modified Fucan 7 in suppressing surgical adhesions, the following double uterine horn (DUH) surgery was performed on both uterine horns of a total of four New Zealand White rabbits. Prior to surgery, the rabbits were weighed and then prepared for surgery with premedication of ketamine and xylazine.

[0243] A 0.33 mg / mL fucoidan solution was prepared in Ringer's lactate solution USP (LRS) and sterilized by filtration. All instruments were sterilized, and a sterile field was maintained during the surgery. The abdomen was cleaned, and access was made via a midline abdominal incision. The uterine horn was located, exposed outside the body, and injured by scraping. The abdominal wall near the scraped uterine horn was also scraped. A minimal amount of fucoidan solution was applied to the injured uterine horn and lateral wall region. The injured uterine horn and abdominal wall were positioned adjacent to each other and fixed with sutures. Before closing the incision, approximately 15 mL / kg of fucoidan solution per rabbit's body weight was applied to the abdominal cavity. Adhesion was evaluated two weeks after the surgery. Three rabbits were evaluated for each prepared fucoidan concentration. The length of the uterine horn adhesion was measured with a ruler. The uterine horn adhesion coverage was calculated using Equation 2.

[0244] Four New Zealand White rabbits were subjected to the same surgical procedure, and instead of fucoidan solution, they were administered approximately 15 mL / kg of the control, Ringer's lactate solution USP (LRS). The control group, administered LRS, had an adhesion coverage rate of 76%, calculated using Equation 2. Table 10 shows the results for Fukan 7 obtained using the method discussed above. The results in the following tables represent the decrease in adhesion coverage rate compared to the control group.

[0245] Table 10 provides the results of the treatment of 8 uterine horns with fucoidan 7.

[0246]

Table 10

[0247] As can be seen from the results in Table 10, fucoidan 7, which was prepared using a known method and has a total non - fucoidan component content of more than 50%, was not effective in treating fibrous adhesions.

[0248] Example 23 Uterine horn fibrous adhesions treated with fucoidan 4 To measure the effectiveness of purified / modified fucoidan 4 in suppressing surgical adhesions, the following double uterine horn (DUH) surgery was performed on both uterine horns of a total of 3 New Zealand White rabbits. Before surgery, the rabbits were weighed, and then prepared for surgery by pre - dosing with ketamine and xylazine.

[0249] A 5 mg / mL fucoidan solution in lactated Ringer's solution USP (LRS) was prepared and sterilized by filtration. All instruments were sterilized, and a sterile field was maintained throughout the surgery. The abdomen was cleaned and accessed by a mid - abdominal incision. The position of the uterine horn was determined, exposed, and abraded to induce injury. The abdominal wall near the abraded uterine horn was also abraded. The damaged uterine horn and abdominal wall were placed adjacent to each other and fixed with sutures. The upper one - third and lower one - third of the myotomy were closed, and a fucoidan solution of 5 mL / kg per rabbit body weight was applied intraperitoneally. The myotomy was temporarily closed, and the fucoidan solution was left in the peritoneal cavity for 30 minutes. The myotomy was opened again, and the peritoneal cavity was washed with 10 mL / kg of LRS. Most of the fluid in the peritoneal cavity was aspirated and removed before closing the incision. Adhesion formation was evaluated 2 weeks after the surgery. The length of the uterine horn adhesion was measured with a ruler. The uterine horn adhesion coverage rate, which is the ratio of the length of the adhesion to the total damaged uterine horn length, was calculated using Equation 2.

[0250] Table 11 shows the results obtained for Fukan 4, a representative example of purified / modified Fukan, using the methods discussed above. The results in the following table are shown as the average adhesion length across the six uterine horns evaluated.

[0251] Table 11 shows the results of treatment of six uterine horns by Fukan 4.

[0252] [Table 11]

[0253] As can be seen from the results in Table 11, purified / modified Fucan can be used to successfully suppress, prevent, eliminate, alleviate, or otherwise treat postoperative uterine horn adhesions.

[0254] Example 24: Uterine horn fibrous adhesions treated with purified / modified Fukan composition To measure the efficacy of a purified / modified fucose composition containing 92% w / w or more of fucose, galactose, sulfate, and counterions in inhibiting surgical adhesions, the following double uterine horn (DUH) surgery was performed on both uterine horns of a total of 20 New Zealand White rabbits. Prior to surgery, the rabbits were weighed and then prepared for surgery with premedication of midazolam and dexmedetomidine.

[0255] Fucoidan solutions at concentrations of 0.02 mg / mL, 0.1 mg / mL, 0.5 mg / mL, or 2.5 mg / mL were prepared in Ringer's lactate solution USP (LRS) and sterilized by filtration. All instruments were sterilized, and a sterile field was maintained throughout the surgery. The abdomen was cleaned, and access was made via a midline abdominal incision. The uterine horn was located, exposed, and scraped to induce damage. The abdominal wall near the scraped uterine horn was also scraped. The damaged uterine horn and abdominal wall were positioned adjacent to each other and fixed with sutures. Before closing the incision, approximately 2 mL / kg of fucoidan solution per rabbit's body weight was applied to the abdominal cavity. Adhesion was evaluated two weeks after the surgery. Five rabbits were treated and evaluated with each prepared fucoidan concentration. The length of the uterine horn adhesion was measured with a ruler. The uterine horn adhesion coverage rate was calculated using Equation 2.

[0256] As a control group, five additional New Zealand White rabbits were subjected to the same surgical procedure, and each was administered approximately 2 mL / kg of control Ringer's lactate solution USP (LRS) instead of fucoidan solution. The control group, which received LRS, had a 100% adhesion coverage rate, as calculated using Equation 2. Table 12 shows the results obtained using the methods discussed above for purified / modified fucoidan compositions at different concentrations and doses (10 uterine horns were treated for each concentration of purified / modified fucoidan composition, for a total of 40 uterine horns). The results are shown as a decrease in adhesion coverage rate compared to the control group.

[0257] [Table 12]

[0258] As can be seen from the results in Table 12, purified / modified Fucan compositions can be used to successfully suppress, prevent, eliminate, alleviate, or otherwise treat postoperative uterine horn adhesions. [Explanation of Symbols]

[0259] 1200 Cation Content Modification System 1202 Inflow Supply Line 1204 Pre-filter 1206 Cation Content Change System Drain Valve 1208 Cation Content Change System Drain Line 1210 Tangential Flow Filtration (TFF) Filter 1212 TFF Supply Line 1214 TFF Inlet Pump 1216 Fukkan Container 1217 TFF Hold Solution Valve 1218 TFF Hold Solution Return Line 1219 TFF Filtration Product Outflow Line 1220 First Dialfiltration Solution Container 1224 First Dialfiltration Solution Valve 1225 First Dialfiltration Solution Supply Line 1230 Second Dialfiltration Solution Container 1234 Second Dialfiltration Solution Valve 1235 Second Dialfiltration Solution Supply Line

[0260] All terms used in this specification are used according to their ordinary meanings, unless the context or definition clearly indicates otherwise. Also, unless explicitly stated otherwise, in this disclosure, the use of "or" includes "and" and vice versa. Non-limiting terms are not to be construed as limiting, unless explicitly stated or the context clearly indicates otherwise (e.g., "including", "having", and "comprising" typically indicate "including without limitation"). Singular forms, such as "a", "an", and "the", include plural references, including in the claims, unless explicitly stated or the context clearly indicates otherwise.

[0261] Unless otherwise specified, adjectives in this specification such as “substantially” and “about” that modify the condition or relational characteristics of one or more features of an embodiment indicate that the condition or characteristic is defined within a permissible range for the implementation of the embodiment for which it is intended.

[0262] The scope of this method, composition, system, etc., includes both the means-plus-function concept and the step-plus-function concept. However, a claim is not interpreted as indicating a "means-plus-function" relationship unless the word "means" is specifically described in the claim, but is interpreted as indicating a "means-plus-function" relationship if the word "means" is specifically described in the claim. Similarly, a claim is not interpreted as indicating a "step-plus-function" relationship unless the word "step" is specifically described in the claim, but is interpreted as indicating a "step-plus-function" relationship if the word "step" is specifically described in the claim.

[0263] From the foregoing, it will be understood that specific embodiments have been considered herein for illustrative purposes, but various modifications may be made without departing from the spirit and scope of the considerations herein. Accordingly, the systems and methods, etc., include, but are not limited to, such modifications and all permutations and combinations of the subject matter described herein, except as provided in the appended claims or other claims that are adequately supported in the considerations and figures herein. Exemplary embodiments of the present invention are described below. <1> A fucane composition comprising at least about 96% w / w fucane and about 4% w / w or less of non-fucan components, compounds, or substances, wherein the fucane comprises fucose, sulfuric acid, and counterions, and may further consist of fucose, sulfuric acid, counterions, galactose, glucose, rhamnose, mannose, xylose, and glucuronic acid. <2> The aforementioned fucose contains approximately 17% w / w or less of counterions, and the total content of fucose, galactose, sulfate, and counterions in the fucose is greater than 96% w / w. <1> The Fukan composition described above. <3> The aforementioned fucane consists of fucose, galactose, sulfate, and counterions. <1> The Fukan composition described above. <4> The aforementioned impurities include water. <1> ~ <3> A Fukan composition as described in any one of the following. <5> The above-mentioned fukan and the above-mentioned impurities, which consist of approximately 3% or less <1> ~ <4> A Fukan composition as described in any one of the following. <6> The above-mentioned fukan and the above-mentioned impurities, which consist of approximately 2% or less <1> ~ <4> A Fukan composition as described in any one of the following. <7> The above-mentioned fukan and the impurities comprising about 1% or less, <1> ~ <4> A Fukan composition as described in any one of the following. <8> The above consists of the above-mentioned fukan and the above-mentioned impurities in an amount of about 0.1% or less, <1> ~ <4> A Fukan composition as described in any one of the following. <9> The fucose content of the aforementioned fucane is more than 25% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <10> The fucose content of the aforementioned fucane is more than 30% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <11> The fucose content of the aforementioned fucane is more than 35% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <12> The fucose content of the aforementioned fucane is more than 40% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <13> The galactose content of the aforementioned Fucan is less than 10% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <14> The galactose content of the aforementioned Fucan is less than 5% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <15> The total counterion content of the aforementioned Fukan is less than 17% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <16> The total counterion content of the aforementioned Fukan is less than 14% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <17> The total counterion content of the aforementioned Fukan is less than 10% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <18> The total counterion content of the aforementioned Fukan is less than 7% w / w. <1> ~ <8> A Fukan composition as described in any one of the following. <19> The aforementioned counterion is a pharmaceutically acceptable counterion. <1> ~ <18> A Fukan composition as described in any one of the following. <20> The pharmaceutically acceptable counterion comprises at least one of aluminum, arginine, benzathine, chloroprocaine, choline, sodium, potassium, lithium, ammonium, ethylenediamine, diethylamine, diethanolamine, ethanolamine, histidine, lysine, N-methylglucamine, meglumine, procaine, triethylamine, zinc, calcium, and magnesium. <19> The Fukan composition described above. <21> The pharmaceutically acceptable counterion comprises at least one of sodium and potassium. <19> The Fukan composition described above. <22> The aforementioned pharmaceutically acceptable counterion essentially consists of at least one of sodium and potassium. <19> The Fukan composition described above. <23> One 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm and packed with a hydroxylated polymethacrylate gel with an effective molecular weight range of approximately 50 kDa to approximately 5,000 kDa, another 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm and packed with a hydroxylated polymethacrylate gel with an effective molecular weight range of approximately 1 kDa to approximately 6,000 kDa, and a 40 mm guard column with an inner diameter of 6 mm and packed with a hydroxylated polymethacrylate gel, wherein the two analytical gel permeation chromatography columns and the one guard column are housed in a column compartment at approximately 30°C; A differential refractive index detector with a temperature of approximately 30°C; 0.6 mL / min 0.1 M sodium nitrate mobile phase flow; and Quantification of the peak molecular weight standard curve, which is essentially composed of the following dextran standards: 1st dextran standard with a peak molecular weight of approximately 2,200 kDa, 2nd dextran standard with a peak molecular weight of approximately 720 kDa to 760 kDa, 3rd dextran standard with a peak molecular weight of approximately 470 kDa to 510 kDa, 4th dextran standard with a peak molecular weight of approximately 370 kDa to 410 kDa, 5th dextran standard with a peak molecular weight of approximately 180 kDa to 220 kDa, and 6th dextran standard with a peak molecular weight of approximately 40 kDa to 55 kDa. When measured using a substantially aqueous gel permeation chromatography configuration, the Fukan has a molecular weight distribution in which at least 60% w / w of the distribution is greater than 100 kDa. <1> ~ <22> A Fukan composition as described in any one of the following. <24> The aforementioned Fukan has a molecular weight distribution in which at least 92% w / w of the distribution is greater than 100 kDa. <23> The Fukan composition described above. <25> The aforementioned Fukan has a molecular weight distribution in which at least 97% w / w of the distribution is greater than 100 kDa. <23> The Fukan composition described above. <26> The aforementioned Fukan has a weight-average molecular weight greater than 100 kDa. <23> The Fukan composition described above. <27> The sulfation level of the aforementioned Fukan is approximately 20% w / w to 60% w / w. <1> ~ <26> A Fukan composition as described in any one of the following. <28> The sulfation level of the aforementioned Fukan is approximately 30% w / w to 55% w / w. <1> ~ <26> A Fukan composition as described in any one of the following. <29> The sulfation level of the aforementioned Fukan is approximately 35% w / w to 52% w / w. <1> ~ <26> A Fukan composition as described in any one of the following. <30> The total carbohydrate content of the aforementioned Fukan is 27% w / w to 80% w / w. <1> ~ <26> A Fukan composition as described in any one of the following. <31> The total content of glucuronic acid, mannose, rhamnose, glucose, and xylose in the aforementioned Fucan is less than approximately 12% w / w as a percentage of the total carbohydrate content. <30> The Fukan composition described above. <32> When dissolved in water at a concentration of 50 mg / mL, it has a viscosity of approximately 4 cP to 50 cP. <1> ~ <31> A Fukan composition as described in any one of the following. <33> When dissolved in water at a concentration of 50 mg / mL, it has a viscosity of approximately 15 cP to 30 cP. <1> ~ <31> A Fukan composition as described in any one of the following. <34> It is a white solid. <1> ~ <33> A Fukan composition as described in any one of the following. <35> When dissolved in water at a concentration of 1 mg / mL to 100 mg / mL, it forms a colorless, transparent solution. <1> ~ <34> A Fukan composition as described in any one of the following. <36> The acetyl content of the aforementioned Fucan is less than approximately 5% w / w. <1> ~ <35> A Fukan composition as described in any one of the following. <37> The acetyl content of the aforementioned Fucan is less than approximately 2% w / w. <1> ~ <35> A Fukan composition as described in any one of the following. <38> When measured by 2D1H-13C heteronuclear multi-quantum coherence in the range of 10-30 ppm in the carbon dimension, at 70°C with a 600 MHz spectrometer equipped with a 5 mm cold probe, suppressing solvent signals, the acetyl content of the fukan is substantially 0% w / w. <1> ~ <35> A Fukan composition as described in any one of the following. <39> <1> ~ <38> A method comprising preparing a Fukan composition as described in any one of the above. <40> <1> ~ <38> A method comprising using any one of the Fukan compositions described. <41> The aforementioned use includes treating fibrous adhesions. <40> Methods used. <42> A therapeutically effective amount in a medically acceptable buffer or diluent <1> ~ <38> A medically acceptable composition comprising the Fukan composition described in any one of the above. <43> A method for treating a condition or disease in an animal, wherein for the purpose of treating the said condition or disease <42> A method comprising selecting a medically acceptable composition as described above, and administering to the animal a therapeutically effective amount of Fucan, comprising about 0.5 mg / kg to 50 mg / kg. <44> A method for treating a condition or disease in an animal, wherein for the purpose of treating the said condition or disease <42> A method comprising selecting a medically acceptable composition as described above, and administering to the animal a therapeutically effective amount of Fucan of about 0.04 mg / kg to 25 mg / kg. <45> The aforementioned amount is approximately 0.2 mg / kg to 10 mg / kg. <43> or <44> Methods used. <46> The aforementioned amount is approximately 1 mg / kg to 5 mg / kg. <43> or <44> Methods used. <47> The aforementioned amount is approximately 1.5 mg / kg to 3 mg / kg. <43> or <44> Methods used. <48> The aforementioned amount is approximately 5 mg / kg to 10 mg / kg. <43> or <44> Methods used. <49> The condition or disease is a fibrous adhesion at a target site in the animal, and the administration includes administering the therapeutically effective amount to the target site. <43> ~ <48> The method described in any one of the following ways. <50> <1> ~ <38> A medical composition comprising approximately 0.02 mg / mL to 100 mg / mL of the Fukan composition described in any one of the above, wherein the medical composition is configured and composed for treating a disease or condition in an animal. <51> The aforementioned Fukan composition contains approximately 0.5 mg / mL to 5 mg / mL <50> The medical composition described above. <52> The Fukan composition contains approximately 2.5 mg / mL <50> The medical composition described above. <53> The aforementioned medical composition is a medical device. <50> ~ <52> A medical composition as described in any one of the following. <54> The medical composition is a liquid medical device. <50> ~ <52> A medical composition as described in any one of the following. <55> The medical composition is a pharmaceutical composition. <50> ~ <52> A medical composition as described in any one of the following. <56> The medical composition is a liquid pharmaceutical composition. <50> ~ <52> A medical composition as described in any one of the following. <57> The aforementioned disease or condition is fibrous adhesion, <50> ~ <56> A medical composition as described in any one of the following. <58> A dose range of approximately 0.01 mL / kg to 15 mL / kg for treating diseases or conditions in animals. <50> ~ <57> Use of any one of the medical compositions described in that document. <59> A dose range of approximately 0.03 mL / kg to 4 mL / kg for treating diseases or conditions in animals. <50> ~ <57> Use of any one of the medical compositions described in that document. <60> A dose range of approximately 0.06 mL / kg to 2 mL / kg for treating diseases or conditions in animals. <50> ~ <57> Use of any one of the medical compositions described in that document. <61> A dose range including approximately 2 mL / kg to 4 mL / kg for treating diseases or conditions in animals. <50> ~ <57> Use of any one of the medical compositions described in that document. <62> A method for treating a selected disease or condition in a patient, comprising identifying a selected target site in the patient that includes the selected disease or condition or is reasonably susceptible to the selected disease or condition, and subsequently applying to the target site in the patient. <50> ~ <57> A method comprising administering any one of the medical compositions described in the following. <63> The aforementioned disease or condition is fibrous adhesion, <62> Methods used. <64> The target site is a surgical site, and the administration is performed at least one of the following: a) after opening the surgical wound at the surgical site, b) during surgery, and c) after closing the surgical wound. <62> or <63> Methods used. <65> The administration is performed after surgery and before the surgical wound is closed. <62> or <63> Methods used. <66> The administration takes less than 3 minutes. <62> or <63> Methods used. <67> The administration takes less than 2 minutes. <62> or <63> Methods used. <68> The administration takes less than one minute. <62> or <63> Methods used. <69> The target site is at least one of a lesion, abrasion, and injury site. <62> ~ <68> The method described in any one of the following ways. <70> The target site is at least one of the following: pelvic cavity, abdominal cavity, dorsal cavity, cranial cavity, spinal cavity, ventral cavity, thoracic cavity, pleural cavity, pericardial cavity, joints, muscles, tendons, and ligaments. <69> Methods used. <71> A method for removing impurities from a starting fukan composition to obtain a purified / modified fukan composition, To provide a starting Fukan composition containing impurities, A reaction mixture is generated by adding a flocculant to the starting Fukan composition. The impurities are agglomerated by heating the reaction mixture to produce agglomerated impurities, and Removing the aforementioned aggregated impurities A method that includes this. <72> Providing the aforementioned starting Fukan composition includes providing the aforementioned starting Fukan composition as a solution. <71> Methods used. <73> The further step is to recover the purified / modified Fukan composition in a solution from which impurities have been reduced. <71> Methods used. <74> The aggregation of the impurities includes heating the reaction mixture at a pressure exceeding atmospheric pressure. <71> Methods used. <75> The aforementioned flocculation aid contains a salt. <71> Methods used. <76> The salts include alkali metals, alkaline earth metals, aluminum and / or ammonium chlorides, bromides, iodides, fluorides, sulfates, sulfites, carbonates, bicarbonates, phosphates, nitrates, nitrites, acetates, citrates, silicates and / or cyanides. <75> Methods used. <77> The aforementioned flocculation aid contains a base. <71> Methods used. <78> The base includes alkali metals, alkaline earth metals, aluminum and / or ammonium hydroxides and / or oxides. <77> Methods used. <79> The impurities to be removed include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. <71> ~ <78> The method described in any one of the following ways. <80> A method for removing impurities from a starting fukan composition to obtain a purified / modified fukan composition, To provide a starting Fukan composition that is a solid, and an extraction medium configured to dissolve impurities but unable to dissolve Fukan, Mixing the starting Fukan composition with the extraction medium to produce a mixture of the purified / modified Fukan composition and the extraction medium, and Separating the purified / modified Fukan composition from the extraction medium. A method that includes this. <81> The further step is to recover the purified / modified Fukan composition, which is in solid form. <80> Methods used. <82> The extraction medium comprises at least one organic solvent having a relative polarity of less than 0.765. <80> Methods used. <83> The organic solvent includes at least one of ethanol, isopropanol, methanol, benzene, diethyl ether, decamethylcyclopentasiloxane, ethyl acetate, butanol, hexane, heptane, heptanol, octanol, and decanol. <82> Methods used. <84> The extraction medium further comprises at least one of a base, a surfactant, and an oxidizing agent. <82> Methods used. <85> Providing the starting Fukan composition in solid form includes precipitating the starting Fukan composition from a solution. <80> Methods used. <86> The impurities to be removed include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. <80> ~ <85> The method described in any one of the following ways. <87> A method for removing impurities from a starting fukan composition to obtain a purified / modified fukan composition, To provide a starting Fukan composition containing impurities, including suspended impurities, in solution. By using an ionic polyvalent impurity precipitating agent to precipitate the impurities from the solution, a mixture of suspended impurities, precipitated impurities, and a supernatant solution is produced, and To separate the suspended impurities and the precipitated impurities from the supernatant solution. A method that includes this. <88> The further step is to recover the supernatant solution containing the purified / modified Fukan composition. <87> Methods used. <89> The aforementioned ionic polyvalent impurity precipitant contains a salt of a divalent or trivalent cation. <87> Methods used. <90> The salts are chlorides, bromides, iodides, fluorides, sulfates, sulfites, carbonates, bicarbonates, phosphates, nitrates, nitrites, acetates, citrates, silicates, and / or cyanides. <89> Methods used. <91> The cation is an alkaline earth metal, zinc, aluminum, copper and / or iron. <89> Methods used. <92> The ionic polyvalent impurity precipitant contains a base with a divalent or trivalent cation. <87> Methods used. <93> The base is a hydroxide and / or oxide of an alkaline earth metal, zinc, aluminum, copper and / or iron. <92> Methods used. <94> Separating the suspended impurities and precipitated impurities from the supernatant solution includes adding a flocculant to the mixture of the suspended impurities, precipitated impurities and the supernatant solution to floccate the suspended impurities and precipitated impurities. <87> ~ <93> The method described in any one of the following ways. <95> The flocculant comprises at least one of the following: potassium aluminum sulfate, sodium aluminum sulfate, aluminum ammonium sulfate, calcium chloride, sodium phosphate, aluminum hydroxide, aluminum chloride, ferric chloride, ferric sulfate, ferrous sulfate, sodium silicate, calcium silicate, calcium phosphate, zinc chloride, calcium carbonate, calcium bicarbonate, potassium sulfate, magnesium phosphate, acrylamide, acrylic acid, aluminum chlorohydrate, polyaluminum chloride, tannin, formaldehyde, melamine, N,N-dimethylaminoethyl acrylate methyl chloride, N,N-dimethylaminoethyl methacrylate methyl chloride quaternary salt, and polydiallyldimethylammonium chloride. <94> Methods used. <96> This further includes maintaining a pH of approximately 7-14. <87> ~ <95> The method described in any one of the following ways. <97> Maintaining the aforementioned pH includes adding a base. <96> Methods used. <98> The impurities to be removed include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. <87> ~ <97> The method described in any one of the following ways. <99> A method for removing impurities from a starting fukan composition to obtain a purified / modified fukan composition, To provide a starting Fukan composition containing impurities, Adjust the pH of the aforementioned starting Fukan composition to approximately 8-14. Adding a cell-disrupting agent configured to dissolve cellular components to the starting Fukan composition to produce a reaction mixture containing the cell-disrupting agent, a biomolecular lysate, and the starting Fukan composition, and To remove the cell-destructive agent and biomolecular lysate from the reaction mixture. A method that includes this. <100> Providing the aforementioned starting Fukan composition includes providing the aforementioned starting Fukan composition which is a solution. <99> Methods used. <101> The further step is to recover the purified / modified Fukan composition in a solution from which impurities have been reduced. <99> Methods used. <102> The cell-destroying agent includes a surfactant. <99> Methods used. <103> The aforementioned surfactant is an anionic surfactant. <102> Methods used. <104> The aforementioned surfactant is a cationic surfactant. <102> Methods used. <105> The aforementioned surfactant is a nonionic surfactant. <102> Methods used. <106> The surfactant comprises at least one of the following: sodium dodecyl sulfate (SDS), benzalkonium chloride, Triton® X100, Triton® X114, Brij® surfactant, Tween® surfactant, sodium deoxycholate, and alkylbenzene sulfonate. <102> Methods used. <107> Removing the cell-destroying agent and biomolecular lysate involves adding a flocculant configured to agglomerate the cell-destroying agent and biomolecular lysate to the reaction mixture. <99> ~ <106> The method described in any one of the following ways. <108> Removing the cell-disrupting agent involves adding a precipitating agent configured to make the cell-disrupting agent insoluble in the reaction mixture to the reaction mixture to generate a precipitate. <99> ~ <106> The method described in any one of the following ways. <109> Removing the biomolecular lysate includes adding a precipitating agent configured to make the biomolecular lysate insoluble in the reaction mixture to the reaction mixture to generate a precipitate. <99> ~ <108> The method described in any one of the following ways. <110> The further step is to add a flocculant configured to cause the precipitate to flocce to the reaction mixture. <108> and <109> The method described in any one of the following ways. <111> The flocculant comprises at least one of the following: potassium aluminum sulfate, sodium aluminum sulfate, aluminum ammonium sulfate, calcium chloride, sodium phosphate, aluminum hydroxide, aluminum chloride, ferric chloride, ferric sulfate, ferrous sulfate, sodium silicate, calcium silicate, calcium phosphate, zinc chloride, calcium carbonate, calcium bicarbonate, potassium sulfate, magnesium phosphate, acrylamide, acrylic acid, aluminum chlorohydrate, polyaluminum chloride, tannin, formaldehyde, melamine, N,N-dimethylaminoethyl acrylate methyl chloride, N,N-dimethylaminoethyl methacrylate methyl chloride quaternary salt, and polydiallyldimethylammonium chloride. <107> ~ <110> The method described in any one of the following ways. <112> Removing the anionic surfactant includes anionic adsorption. <103> Methods used. <113> Removing the aforementioned cationic surfactant includes cation adsorption. <104> Methods used. <114> Removing the nonionic surfactant involves micelle phase separation. <105> Methods used. <115> Removing the aforementioned surfactant includes hydrophobic adsorption. <102> Methods used. <116> Removing the aforementioned surfactant means Diluting the reaction mixture until the concentration of the surfactant falls below a predetermined concentration, and The reaction mixture containing the surfactant is subjected to dialysis filtration using a tangential flow filter whose molecular weight cutoff exceeds the maximum molecular weight of the surfactant. including, <102> Methods used. <117> The further step involves adding a chelating agent to the reaction mixture after providing the starting Fukan composition and before removing the cell-disrupting agent. <99> ~ <116> The method described in any one of the following ways. <118> The chelating agent comprises ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin and / or citric acid. <117> Methods used. <119> The further step involves adding an oxidant-quenching agent to the reaction mixture before removing the cell-destroying agent to quench the reaction of the oxidant in the reaction mixture. <99> ~ <118> The method described in any one of the following ways. <120> The reaction mixture further comprises adding a bacteriostatic agent after providing the starting Fukan composition and before removing the cell-disrupting agent. <99> ~ <119> The method described in any one of the following ways. <121> The bacteriostatic agent comprises sodium sulfite, ethylenediaminetetraacetic acid (EDTA), benzalkonium chloride, ethanol and / or thiourea. <120> Methods used. <122> The impurities to be removed include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. <99> ~ <121> The method described in any one of the following ways. <123> A method for removing impurities from a starting fukan composition to obtain a purified / modified fukan composition, To provide a starting Fukan composition containing impurities in an aqueous starting solution, The aqueous starting solution is mixed with an organic solvent to produce an aqueous-organic phase mixture, and The aqueous phase-organic phase mixture is separated to obtain an aqueous portion and an organic portion. A method that includes this. <124> The further step is to recover the aqueous portion containing the purified / modified Fucan composition. <123> Methods used. <125> The organic solvent includes at least one organic solvent having a relative polarity of less than 0.765. <123> Methods used. <126> The organic solvent comprises at least one of ethanol, isopropanol, methanol, benzene, decamethylcyclopentasiloxane, ethyl acetate, hexane, heptanol, octanol, decanol, heptane, isobutyl acetate, anisole, isopropyl acetate, 1-butanol, butyl acetate, methyl isobutyl ketone, pentane, 1-pentanol, ethyl ether, and propyl acetate. <123> ~ <125> The method described in any one of the following ways. <127> The impurities to be removed include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. <123> ~ <126> The method described in any one of the following ways. <128> A method for changing the cation content of a starting Fukan composition, To provide the starting Fukan composition in the starting solution, and The starting solution is dialy filtered through a tangential flow filtration filter using a chelating agent solution that passes through a tangential flow filtration filter to produce a retained Fukan composition. A method that includes this. <129> The chelating agent comprises at least one of ethylenediaminetetraacetic acid (EDTA), 2,3-dimercapto-1-propanol, ethylenediamine, porfin, and citric acid. <128> Methods used. <130> The cation content of the retaining fukan composition is essentially composed of sodium and / or potassium. <128> Methods used. <131> A method for removing impurities from a starting fukan composition to obtain a purified / modified fukan composition, To provide a starting Fukan composition containing impurities, The starting Fukan composition is subjected to an appropriate pressure exceeding 70 bar and an appropriate temperature exceeding 30°C in a supercritical extraction apparatus. Filling a supercritical fluid into a supercritical extraction apparatus to remove impurities from the supercritical fluid, and The supercritical fluid containing the extracted impurities is removed after a predetermined time. A method that includes this. <132> The further step is to recover the purified / modified Fukan composition remaining in the supercritical extraction apparatus. <131> Methods used. <133> The aforementioned pressure is approximately 70 bar to approximately 2000 bar. <131> Methods used. <134> The aforementioned temperature ranges from approximately 30°C to approximately 300°C. <131> Methods used. <135> The aforementioned starting Fukan composition is a liquid. <131> Methods used. <136> The aforementioned starting Fukan composition is a solid. <131> Methods used. <137> The supercritical fluid comprises at least one of carbon dioxide, ethanol, ethane, hydrochloric acid, hydrofluoric acid, sulfuric acid, and nitric acid. <131> Methods used. <138> The aforementioned predetermined time ranges from approximately 5 minutes to 50 hours. <131> Methods used. <139> The impurities to be removed include at least one of the following: particulate matter, lipids, fatty acids, phlorotannins, laminarin, alginates, proteins, Maillard reaction products, fucoxanthin, chlorophyll, bacteria, cellular components, and DNA. <131> ~ <138> The method described in any one of the following ways.

Claims

1. A fucane composition comprising at least 96% w / w fucane and 4% w / w or less of non-fucan components, compounds, or substances, wherein the fucane comprises (i) fucose and sulfuric acid, (ii) having a sulfation level of 20% w / w to 60% w / w, and (iii) containing 10% w / w or more and less than 17% w / w of counterions, and in addition these may further consist of galactose, glucose, rhamnose, mannose, xylose, and glucuronic acid, wherein the counterions are counterions to the sulfate group -SO42- contained in the fucane.

2. The Fukan composition according to claim 1, wherein the non-Fukan component contains water.

3. The Fukan composition according to claim 1 or claim 2, comprising the Fukan and 3% or less of the non-Fukan component.

4. The Fukan composition according to claim 1 or claim 2, comprising the Fukan and 2% or less of the non-Fukan component.

5. The Fukan composition according to claim 1 or claim 2, comprising the Fukan and 1% or less of the non-Fukan component.

6. The Fukan composition according to claim 1 or claim 2, comprising the Fukan and 0.1% or less of the non-Fukan component.

7. The fucose composition according to any one of claims 1 to 6, wherein the fucose content of the fucose is more than 25% w / w.

8. The fukan composition according to any one of claims 1 to 7, wherein the galactose content of the fukan is less than 10% w / w.

9. The Fukan composition according to any one of claims 1 to 8, wherein the counterion is a pharmaceutically acceptable counterion.

10. One 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm and packed with a hydroxylated polymethacrylate gel having an effective molecular weight range of 50 kDa to 5,000 kDa, another 300 mm analytical gel permeation chromatography column with an inner diameter of 7.8 mm and packed with a hydroxylated polymethacrylate gel having an effective molecular weight range of 1 kDa to 6,000 kDa, and a 40 mm guard column with an inner diameter of 6 mm and packed with a hydroxylated polymethacrylate gel, wherein the two analytical gel permeation chromatography columns and the one guard column are housed in a column compartment at 30°C; 30°C differential refractive index detector; A 0.1 M sodium nitrate mobile phase flow of 0.6 mL / min; and Quantification of the peak molecular weight standard curve consisting of a first dextran standard with a peak molecular weight of 2,200 kDa, a second dextran standard with a peak molecular weight of 720 kDa to 760 kDa, a third dextran standard with a peak molecular weight of 470 kDa to 510 kDa, a fourth dextran standard with a peak molecular weight of 370 kDa to 410 kDa, a fifth dextran standard with a peak molecular weight of 180 kDa to 220 kDa, and a sixth dextran standard with a peak molecular weight of 40 kDa to 55 kDa. The Fukan composition according to any one of claims 1 to 9, wherein, when measured using an aqueous gel permeation chromatography configuration comprising the above, the Fukan has a molecular weight distribution in which at least 60% of the distribution w / w is greater than 100 kDa.

11. The Fukan composition according to claim 10, wherein the Fukan has a molecular weight distribution in which at least 92% w / w of the distribution is greater than 100 kDa.

12. The fukan composition according to claim 10, wherein the fukan has a weight-average molecular weight greater than 100 kDa.

13. The Fukan composition according to any one of claims 1 to 12, wherein the total carbohydrate content of the Fukan is 27% w / w to 80% w / w.

14. The Fukan composition according to claim 13, wherein the total content of glucuronic acid, mannose, rhamnose, glucose, and xylose in Fukan is less than 12% w / w as a percentage of the total carbohydrate content.

15. The Fukan composition according to any one of claims 1 to 14, having a viscosity of 4 mPa·s to 50 mPa·s (4 cP to 50 cP) when dissolved in water at a concentration of 50 mg / mL.

16. The Fukan composition according to any one of claims 1 to 15, which is a white solid.

17. The Fukan composition according to any one of claims 1 to 16, which forms a colorless and transparent solution when dissolved in water at a concentration of 1 mg / mL to 100 mg / mL.

18. The fukan composition according to any one of claims 1 to 17, wherein the acetyl content of the fukan is less than 5% w / w.

19. Using a 600 MHz spectrometer equipped with a 5 mm cold probe, 2D scanning was performed at 70°C, suppressing solvent signals, with 256 to 512 scans per run, increasing by 8 each time, in the range of 10 to 30 ppm in the carbon dimension. １ H- １３ The fukan composition according to any one of claims 1 to 18, wherein the acetyl content of the fukan is 0% w / w when measured by C-henuclear polyquantum coherence.

20. A medically acceptable composition for the treatment of fibrous adhesions, comprising a therapeutically effective amount of the Fukan composition according to any one of claims 1 to 19.

21. A medical composition for treating fibrous adhesions in animals other than humans, comprising 0.02 mg / mL to 100 mg / mL of the Fukan composition described in any one of claims 1 to 19, and being a liquid pharmaceutical composition.

22. The medical composition according to claim 21, wherein the medical composition is a liquid medical device.

23. The medical composition according to claim 21, wherein the Fukan composition is dissolved in a physiological saline solution.

24. Use of the medical composition according to any one of claims 21 to 23, in a dose range of 0.01 mL / kg to 15 mL / kg, for treating fibrous adhesions in animals other than humans.

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