BR112020010887B1 - PENTOSAN POLYSULFATE, MEDICINE, PH BUFFERING AGENT, AND, USE OF PENTOSAN POLYSULFATE, A PHARMACEUTICALLY ACCEPTABLE SALT THEREOF OR A PHARMACEUTICALLY ACCEPTABLE SOLVATE OF PENTOSAN POLYSULFATE OR A PHARMACEUTICALLY ACCEPTABLE SALT THEREOF - Google Patents

Abstract

The present invention provides a pentosan polysulfate having a uronic acid content of 7.0 mass % to 15.0 mass % and an acetyl group content of 0 mass % to 2.0 mass %; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. Pentosan polysulfate, the pharmaceutically acceptable salt thereof, or the pharmaceutically acceptable solvate of pentosan polysulfate or the pharmaceutically acceptable salt thereof are useful as an active ingredient of a medicament for preventing and/or treating a disease caused by abnormal improvement in the function of FGF-2, and as a pH buffering agent.

Description

Technical Field

[001] The present invention relates to a pentosan polysulfate, and a medicine comprising pentosan polysulfate.

Fundamentals of the Technique

[002] A basic fibroblast growth factor (FGF-2 or b-FGF) is known to be involved in diseases associated with abnormal angiogenesis, such as tumors and arthritis (Patent Literature (PTL) 1). FGF-2 is a heparin-binding growth factor that binds to the FGF-2 receptor of cells by binding to heparan sulfate.

[003] Pentosan polysulfate is known as one of the substances that inactivates FGF-2. Pentosan polysulfate has been reported to inhibit angiogenesis etc. (Non-Patent Literature (NPL) 1 to Non-Patent Literature (NPL) 3). Pentosan polysulfate is considered to bind to FGF-2 and thus prevent FGF-2 from binding to heparan sulfate.

[004] It has also been reported that pentosan polysulfate actually inhibits the growth of tumors (Patent Literature (PTL) 2 and Non-Patent Literature (NPL) 4).

[005] Pentosan polysulfate is produced by chemical sulfation of xylan obtained from hardwoods (e.g. beech). Pentosan polysulfate is composed of a sulfated linear polysaccharide in which β-D-silopyranose is linearly linked; and has 4-O-methylglucuronic acid, that is, uronic acid, for every 10 units of silopyranose (Patent Literature (PTL) 3 and Patent Literature (PTL) 4). Patent literature (PTL) 5 discloses that pentosan polysulfate having a uronic acid content of 4.3 to 6% was obtained by a method comprising fractionating commercially available pentosan polysulfate (SP-54) to obtain a low weight pentosan polysulfate molecular.

Citation list Patent literature

[006] PTL 1: WO2013/186857 PTL 2: JPH3-20225a PTL 3: WO2010/000013 PTL 4: JP2009-532467A PTL 5: JPS61-197601a

Non-patent literature

[007] NPL 1: Gonzalez et al., Biol. Pharm. Bull., 2001; 24; two; 151 154 NPL 2: S. Swain et al., Annals of the New York Academy of Sciences, 1993; 698; 63-67 NPL 3: G. Zugmaier et al., Annals of the New York Academy of Sciences, 1999; 886; 243-248 NPL 4: G. Zugmaier et al., Journal of the National Cancer Institute, 1992; 84; 22; 1716-1724

Summary of the Invention Technical problem

[008] An object of the present invention is to provide a new pentosan polysulfate that has preferable activity for pharmaceutical application, or application as a pH buffering agent.

Means to Solve the Problem

[009] As a result of the intensive study to solve the above problem, the present inventors have verified a new pentosan polysulfate that has a high inhibitory activity to inhibit the binding between FGF-2 and heparan sulfate, compared to conventional pentosan polysulfate. The inventors have further discovered that this pentosan polysulfate can also function as a pH buffering agent. The present invention was made based on these findings.

[0010] Specifically, the present invention provides the following [1] to [13].

[0011] [1] A pentosan polysulfate that has a uronic acid content of 7.0% by mass to 15.0% by mass, and an acetyl group content of 0% by mass to 2.0% by mass; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0012] [2] The pentosan polysulfate according to [1], wherein the pentosan polysulfate has a uronic acid content of 7.5% by mass to 13.0% by mass; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0013] [3] The pentosan polysulfate according to [1] or [2], wherein the pentosan polysulfate has a weight average molecular weight of 5,000 or less; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0014] [4] The pentosan polysulfate according to [3], wherein the pentosan polysulfate has an acetyl group content of 0 to 0.3% by mass; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0015] [5] The pentosan polysulfate according to any one of [1] to [4], wherein the pentosan polysulfate has a structure represented by Formula II: Formula II wherein R each independently represents a hydrogen atom, -COCH3, or -SO3X1, and at least one R in the molecule is -SO3X1, wherein X1 represents a hydrogen atom or a monovalent or divalent metal; X represents a hydrogen atom or a monovalent or divalent metal; and n1 and n2 each independently represent an integer of 0 or more and 30 or less, and at least one of n1 and n2 is an integer of 1 or more; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0016] [6] The pentosan polysulfate according to [5], where each R independently represents a hydrogen atom or -SO3X; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0017] [7] The pentosan polysulfate according to [5] or [6], where X is sodium; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0018] [8] A medicament comprising as an active ingredient pentosan polysulfate according to any one of [1] to [7]; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof.

[0019] [9] The medicine according to [8], which is for use in the prevention and/or treatment of a disease caused by abnormal increase in FGF-2 function.

[0020] [10] The medicine according to [9], wherein the disease caused by abnormal increase in FGF-2 function is cancer, autoimmune disease, allergic disease, inflammatory disease, cardiac dysplasia, vascular dysplasia, or skeletal dysplasia .

[0021] [11] The medicament according to claim 9, which is for use in the prevention and/or treatment of cystitis or arthritis.

[0022] [12] The medicament according to any one of [8] to [11], which is an injectable formulation.

[0023] [13] A pH buffering agent comprising pentosan polysulfate according to any one of [1] to [7]; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or a pharmaceutically acceptable salt thereof.

[0024] From another point of view, the present invention provides: a method for preventing and/or treating a disease caused by an abnormal increase in FGF-2 function, comprising administering an effective amount of pentosan polysulfate according to any one of [ 1] to [7], a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate of pentosan polysulfate or a pharmaceutically acceptable salt thereof to a human or an animal; the use of the pentosan polysulfate according to any one of [1] to [7], a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate of the pentosan polysulfate, or a pharmaceutically acceptable salt thereof, to produce a medicament for preventing and/or treat a disease caused by abnormal increase in FGF-2 function; the use of pentosan polysulfate according to any one of [1] to [7], a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate of pentosan polysulfate, or a pharmaceutically acceptable salt thereof, to prevent and/or treat a disease caused by abnormal increase in FGF-2 function; and the pentosan polysulfate according to any one of [1] to [7], a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate of the pentosan polysulfate or a pharmaceutically acceptable salt thereof, for use as a medicament for preventing and /or treat a disease caused by abnormal increase in FGF-2 function.

Advantageous Effects of the Invention

[0025] The present invention provides a pentosan polysulfate that has a high inhibitory activity to inhibit the binding between FGF-2 and heparan sulfate. The pentosan polysulfate of the present invention is useful as a medicament for preventing and/or treating a disease caused by abnormal increase in FGF-2 function, such as cancer or arthritis. Additionally, the pentosan polysulfate of the present invention can also be used as a pH buffering agent.

Brief Description of the Drawings

[0026] Figure 1 is a graph showing the effect of the uronic acid content of pentosan polysulfate on the inhibitory activity to inhibit the binding between FGF-2 and heparan sulfate.

[0027] Figure 2 is a graph showing the effect of the acetyl group content of pentosan polysulfate on the inhibitory activity to inhibit the binding between FGF-2 and heparan sulfate.

[0028] Figure 3 is a graph showing the relationship between the uronic acid content of pentosan polysulfate and the amount (mL) of a 0.01N aqueous hydrochloric acid solution required to adjust the pH from pH 6 to pH 4 in the titration of pentosan polysulfate solution 100 mg/100 mL.

Description of Modalities

[0029] The present invention is described in detail below. The characteristics of the constituent can be described below based on typical embodiments and specific examples; however, the present invention is not limited to such embodiments.

[0030] In the present specification, “comprising . . . as an active ingredient” means containing as a main active ingredient, and containing in such an amount that an effect is shown.

[0031] The phrase “prevention and/or treatment” means “prevention,” “treatment,” or “prevention and treatment.” For example, the “medicine to prevent and/or treat” may function only as a prophylactic agent, or as a therapeutic agent; or it may have functions as both a prophylactic agent and a therapeutic agent.

Pentosan polysulfate

[0032] Pentosan polysulfate is a compound obtained by sulfation of at least one hydroxyl group of xylo-oligosaccharide. In the present specification, pentosan polysulfate includes pentosan polysulfate salts, pentosan polysulfate solvates, and pentosan polysulfate salt solvates. Pentosan polysulfate salts are preferably pharmaceutically acceptable salts, and examples include sodium pentosan polysulfate, potassium pentosan polysulfate, calcium pentosan polysulfate, and the like. The solvates are preferably pharmaceutically acceptable solvates. Examples of solvents include water.

[0033] Pentosan polysulfate has a structure represented by Formula II. Pentosan polysulfate may contain one structure represented by Formula II, or may contain two or more structures represented by Formula II. When pentosan polysulfate contains two or more structures represented by Formula II, the structure represented by Formula II is a structure representing a pentosan polysulfate repeating unit.

[0034] In Formula II, R each independently represents a hydrogen atom, -COCH3, or -SO3X1, and at least one R in the molecule is -SO3X1, wherein X1 represents a hydrogen atom or a monovalent or divalent metal, and X1 is preferably a hydrogen, sodium, potassium, or calcium atom, more preferably sodium, potassium, or calcium, and particularly preferably sodium; X is a hydrogen atom or a monovalent or divalent metal, and X is preferably sodium, potassium, or calcium, and is particularly preferably sodium; and n1 and n2 each independently represent an integer of 0 or more and 12 or less, and at least one of n1 and n2 is an integer of 1 or more.

[0035] In Formula II, n1 + n2 is preferably from 1 to 10, more preferably from 2 to 8, and even more preferably from 3 to 6.

[0036] The portion that is an end of the structure represented by Formula II and that does not connect to a structure represented by Formula II can be -OR. That is, -OR can be connected to the left terminal (n1 side) of Formula II, while -R can be connected to the right terminal (n2 side) of Formula II. It is particularly preferred that -OR1X connects to the left terminal (n1 side) of Formula II, and -R1X connects to the right terminal (n2 side) of Formula II. In Formula II, R1X is a hydrogen atom or -SO3X1; X1 is a hydrogen atom or a monovalent or divalent metal; and X1 is preferably a hydrogen, sodium, potassium, or calcium atom, more preferably sodium, potassium, or calcium, and particularly preferably sodium.

[0037] In the previous formula, X is preferably a monovalent or divalent metal. A pharmaceutically acceptable salt of pentosan polysulfate is preferred. For example, X is preferably sodium, potassium or calcium. In this case, the pentosan polysulfate salt is pentosan sodium polysulfate, pentosan polysulfate potassium, or pentosan polysulfate calcium. Among them, the pentosan polysulfate salt is particularly preferably sodium pentosan polysulfate.

[0038] The pentosan polysulfate of the present invention has a uronic acid content of 7.0% by mass to 15.0% by mass. The pentosan polysulfate of the present invention preferably has a uronic acid content of 7.5 mass % to 14.0 mass %, and more preferably 7.7 mass % to 13.0 mass %. The above ratio does not have to be met by a single molecule, but can be met by pentosan polysulfate as a complete mixture of individual molecules.

[0039] The pentosan polysulfate of the present invention can be a mixture of molecules represented by Formula II that are different from each other in values n1 and n2, in the type of substituent R, and/or in the degree of substitution.

[0040] Pentosan polysulfate has a structure obtained by sulfation of a xylo-oligosaccharide. The pentosan polysulfate of the present invention is preferably obtained by sulfation of an acidic xylooligosaccharide. Among the xylo-oligosaccharides that have the structure obtained by sulfation of a xylo-oligosaccharide, neutral xylo-oligosaccharides are xylo-oligosaccharides that do not contain uronic acid. Acidic xylooligosaccharides are xylooligosaccharides in which at least one uronic acid is linked to at least one of the xylose units in a xylooligosaccharide molecule. That is, acidic xylooligosaccharides have at least one uronic acid residue as a side chain per xylooligosaccharide molecule. The number of uronic acid residues contained per xylooligosaccharide molecule can be measured by the carbazole sulfuric acid method, or the colorimetric method using sodium tetraborate. The uronic acid content (% by mass) of pentosan polysulfate refers to a calculated value of the number of uronic acid residues in a predetermined amount of pentosan polysulfate obtained by the carbazole sulfuric acid method, as described in the Examples.

[0041] The sulfur content of the pentosan polysulfate of the present invention is preferably 10.0% by mass or more, more preferably 12.0% by mass or more, even more preferably 15.5% by mass or more, and particularly preferably 16.5% by mass or more. The sulfur content of the pentosan polysulfate is preferably 20.0% by mass or less. Here, the sulfur content of pentosan polysulfate is a value determined according to the oxygen flask combustion method described in the Japanese Pharmacopoeia.

[0042] The known pentosan polysulfate is considered to contain a certain amount of xylose units to which one or more acetyl groups (-COCH3) are attached to the uronic acid residues (see, for example, WO2014/114723). In contrast, the pentosan polysulfate of the present invention preferably has an acetyl group content of 0 to 2.0% by mass, more preferably 0 to 1.0% by mass, even more preferably 0 to 0.4% by mass, even more preferably 0 to 0.3% by mass, and particularly preferably substantially 0% by mass. To obtain pentosan polysulfate having an acetyl group content of 0 to 2.0% by mass, the pentosan polysulfate of the present invention is preferably produced by means of a deacetylation step described below.

[0043] The acetyl group content of pentosan polysulfate can be calculated from the integral ratio of the peaks in H NMR measurement. Specifically, first, the HNMR measurement is performed using a HNMR measurement solution containing a specific amount of pentosan polysulfate and a specific amount of an internal standard substance. By comparing the peak for acetyl group to the peak for a specific group of the internal standard substance in the spectrum obtained to obtain an integral ratio thereof, the molar quantity of acetyl groups in the solution is obtained. The molar amount of acetyl groups is then multiplied by 43; and the value obtained is divided by the average molecular weight obtained separately, in order to obtain the mass% of the acetyl groups.

[0044] The weight average molecular weight (Mw) of the pentosan polysulfate of the present invention is not particularly limited; and may be, for example, 5,000 or less, 4,000 or less, 3,900 or less, or 3,800 or less, or 3,750 or less. In this case, the lower limit of the weight average molecular weight (Mw) of the pentosan polysulfate is preferably 1,000.

[0045] The number average molecular weight (Mn) of pentosan polysulfate is not particularly limited; and may be, for example, 5,000 or less, 4,000 or less, 3,900 or less, 3,800 or less, or 3,750 or less. In this case, the lower limit of the number average molecular weight (Mn) of the pentosan polysulfate is preferably 300.

[0046] The weight average molecular weight (Mw) and the number average molecular weight (Mn) of the pentosan polysulfate of the present invention can be measured by GPC (gel permeation chromatography). As the GPC column, a YMC-Pack Diol-300 and YMC-Pack Diol-60 (both manufactured by YMC) can be used connected to each other. The GPC conditions could be, for example, the following conditions.

[0047] Eluent: 25 mM potassium dihydrogen phosphate / 25 mM dipotassium hydrogen phosphate / 50 mM potassium chloride Flow rate: 0.7 mL/min Measuring temperature: 40°C Detector: refractive index detector

[0048] The degree of dispersion of pentosan polysulfate is preferably 1.00 or more and 1.6 or less, more preferably 1.00 or more and 1.5 or less. The degree of dispersion of the pentosan polysulfate is also preferably 1.00 or more and 1.4 or less. The degree of dispersion (D) of pentosan polysulfate is calculated by the following formula. Degree of dispersion (D) = Weight average molecular weight (Mw)/Number average molecular weight (Mn)

[0049] The pentosan polysulfate obtained by the production method of the present invention described below has high purity, and tends to have a narrow molecular weight distribution. The pentosan polysulfate obtained by the production method of the present invention has excellent stability quality.

Application of Pentosan Polysulfate: Medicine

[0050] The pentosan polysulfate of the present invention can be used for applications such as components of medicines, foods, cosmetics and other compositions.

[0051] The pentosan polysulfate of the present invention is particularly useful as an active ingredient of a medicine.

[0052] Examples of medications include medications for preventing and/or treating a disease caused by an abnormal increase in FGF-2 function.

[0053] FGF-2 (basic fibroblast growth factor) is one of the growth factors, and is secreted from various cells. FGF-2 is deeply involved in cell proliferation and differentiation at developmental stages, and is highly expressed during tissue repair in vivo. Additionally, FGF-2 is involved in abnormal angiogenesis, and has potent growth and migration-promoting effects on vascular endothelial cells. FGF-2, which has these functions, is known to be involved in diseases such as tumors. It was also clarified that FGF-2, which promotes angiogenesis and bone destruction, is a fundamental molecule involved in the pathology in chronic rheumatoid arthritis. Particularly, high serum FGF-2 concentration has been reported in tumor with many blood vessels, such as kidney cancer. FGF-2 is also present in several other tumors, such as prostate cancer, breast cancer and lung cancer.

[0054] FGF-2 binds to an FGF receptor (FGFR), which induces the expression of several cytokines and receptor genes. FGF-2 has a heparin-binding region and binds to heparin and heparan sulfate. It is considered that upon binding to FGFR, FGF-2 secreted from a cell is first bound to heparan sulfate from an extracellular matrix, concentrated and protected from the protease. In this way, the activity of inhibiting the binding between FGF-2 and heparan sulfate can be an index to determine the effect of prevention and/or treatment of a disease caused by an abnormal increase in FGF-2 function.

[0055] As shown in the Examples, pentosan polysulfate has an activity of inhibiting the binding between FGF-2 and heparan sulfate; and this inhibitory activity is high when the pentosan polysulfate has a uronic acid content of 7.0 to 15.0% by mass, and an acetyl group content of 0 to 2.0% by mass. Therefore, the pentosan polysulfate of the present invention is particularly useful for preventing and/or treating diseases caused by abnormal increase in FGF-2 function.

[0056] Specific examples of abnormal increase in FGF-2 function include abnormal angiogenesis by FGF-2. Abnormal increase in FGF-2 function can be determined, for example, by using an increase in serum FGF-2 concentration as an index.

[0057] Specific examples of diseases caused by abnormal increase in FGF-2 function include tumors, chronic inflammation, such as arthritis, autoimmune disease, allergic disease, inflammatory disease, such as cystitis, cardiac dysplasia, vascular dysplasia, skeletal dysplasia, psoriasis , age-related macular degeneration, periodontal disease, scleroderma, neovascular glaucoma, and the like.

[0058] The pentosan polysulfate of the present invention is also useful as an active ingredient of a medicament for preventing and/or treating cystitis, particularly interstitial cystitis.

[0059] The dosage form of the medicine of the present invention is not particularly limited, and the medicine can be administered orally or parenterally. Preferably, the medicament may be administered parenterally by injection or intravenous infusion.

[0060] The medicine of the present invention may consist only of pentosan polysulfate, which is an active ingredient. Preferably, however, one or more appropriate pharmacologically and pharmaceutically acceptable additives may be added to the pentosan polysulfate to provide a medicament in a form well known to those skilled in the art.

[0061] Examples of pharmacologically and pharmaceutically acceptable additives include excipients, disintegrants or disintegration aids, binders, lubricants, coating agents, pigments, diluents, bases, solubilizers or dissolution aids, isotonizing agents, buffers, pH adjusters, stabilizers, propellants, adhesives and the like.

[0062] Examples of pharmaceutical preparations suitable for oral administration include tablets, capsules, powders, fine granules, granules, liquids, syrups and the like. Examples of formulations for parenteral administration include injectable formulations, drip infusions, suppositories, inhalants, patches and the like. To prepare formulations suitable for oral administration, transdermal administration, or transmucosal administration, for example, the following pharmacologically and pharmaceutically acceptable additives may be added. Examples of additives include excipients such as glucose, lactose, D-mannitol, starch, and crystalline cellulose; disintegrants or disintegration aids, such as carboxymethylcellulose, starch, and calcium carboxymethylcellulose; binders, such as hydroxypropylcellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, and gelatin; lubricants such as magnesium stearate and talc; coating agents such as hydroxypropylmethylcellulose, sucrose, polyethylene glycol, and titanium oxide; and bases, such as petrolatum, liquid paraffin, polyethylene glycol, gelatin, kaolin, glycerin, purified water, and hard fat. Other examples include propellants such as chlorofluorocarbon, diethyl ether, and compressed gas; adhesives, such as poly(sodium acrylate), poly(vinyl alcohol), methylcellulose, polyisobutylene, and polybutene; and base fabric, such as fabric and cotton and plastic sheets. Pharmaceutical preparations can be formed using such additives for pharmaceutical preparations.

[0063] To prepare pharmaceutical preparations suitable for injection or infusion, for example, the following additives for pharmaceutical preparations can be added. Examples of useful additives include solubilizing agents or solubilization aids that can form ready-to-use aqueous injectable formulations, such as distilled water for injection, saline, and propylene glycol; isotonizing agents, such as glucose, sodium chloride, D-mannitol, and glycerin; buffers, such as phosphates (e.g., disodium hydrogen phosphate and sodium dihydrogen phosphate), citrate, and acetate; and pH regulators, such as inorganic acids, organic acids, inorganic bases, and organic bases.

[0064] As described later, the pentosan polysulfate of the present invention has a greater pH buffering capacity than pentosan polysulfate which has a lower uronic acid content. Therefore, when the pH needs to be adjusted, for example, when the pentosan polysulfate of the present invention is to be provided in the form of a liquid preparation, an injectable formulation, a drip infusion, or the like, it is unnecessary to use a pH adjuster. or the amount of pH adjuster used can be reduced.

[0065] The dosage of the medicine of the present invention is not particularly limited, and can be appropriately selected according to the form of administration; the age, severity of the disease, symptoms, and body weight of the patient; and other conditions. For example, when administered intravenously, subcutaneously, or intramuscularly, the medicament may be administered in an amount of 0.1 to 20 mg/kg, preferably 0.2 to 10 mg/kg, per day for an adult, in terms of the ingredient. active.

Use of Pentosan Polysulfate: Anticoagulant

[0066] The pentosan polysulfate of the present invention can be used as an active ingredient of an anticoagulant.

[0067] The anticoagulants containing the pentosan polysulfate of the present invention can be provided not only as medicines, but also as surface treatment agents for medical devices or medical materials. For example, such anticoagulants can be used as surface treatment agents for implantable artificial organs, artificial blood vessels, catheters, stents, blood bags, contact lenses, intraocular lenses and surgical auxiliary instruments. Examples of methods for immobilizing the pharmaceutical composition on the surface of a medical device or a medical material include a method comprising contacting the pharmaceutical composition with the medical device or the medical material, and irradiating the contact portion with radiation.

Use of Pentosan Polysulfate: pH Buffer

[0068] As shown in the Examples, the pentosan polysulfate of the present invention has a greater pH buffering capacity than the pentosan polysulfate which has a lower uronic acid content. In this way, the pentosan polysulfate of the invention can be used as a pH buffering agent. The pentosan polysulfate of the present invention exhibits buffering action to maintain the pH in the range of pH 4 to pH 6. For example, an injectable formulation that has a pH less than 4 causes pain to a patient. The pentosan polysulfate of the present invention is an active ingredient of a medicine, and can also function as a pH adjuster in an injectable formulation. The pentosan polysulfate of the present invention can be used for foods, medicines, and any other compositions whose pH must be maintained in the range of pH 4 to pH 6 from the point of view of stabilization and prevention of degradation.

[0069] When a composition, such as an aqueous solution, contains the pentosan polysulfate of the present invention as a pH buffering agent, the concentration of pentosan polysulfate is preferably 10 to 500 mg/mL, and more preferably 50 to 300 mg/mL .

Method for producing Pentosan Polysulfate

[0070] The pentosan polysulfate of the present invention can be obtained, for example, by a method for producing pentosan polysulfate, the method comprising Step I of obtaining an acidic xylo-oligosaccharide from a plant-derived raw material, and Step II of obtaining pentosan polysulfate from the acidic xylo-oligosaccharide; and further comprising a deacetylation step. In this method, Step I includes a step of depolymerizing a plant-derived raw material. Since the method comprises the step of depolymerizing a plant-derived raw material and the step of sulfation in that order, the method can effectively produce pentosan polysulfate. The method for producing pentosan polysulfate may additionally include a deacetylation step. By including a deacetylation step, the method can produce a pentosan polysulfate that has a low acetyl group content.

Plant-derived raw material

[0071] Acidic xylo-oligosaccharide can be obtained by depolymerization of a plant-derived raw material. Examples of plant-derived raw materials include wood-derived raw materials, seed-derived raw materials, grain-derived raw materials, fruit-derived raw materials, and the like. Additionally, examples of plant-derived raw materials that can be used include cottons, such as cotton linters and cotton fiber; herbaceous plants, such as kenaf, hemp, ramie, and rice straw; and the like. Like the plant-derived raw material, the previously mentioned raw materials derived from various sources can also be used in combination.

[0072] Among these, wood-derived raw materials are preferably used as the plant-derived raw material. Examples of useful raw materials derived from wood include wooden raw materials such as softwoods and hardwoods. The wood-derived raw material is preferably at least one selected from softwoods and hardwoods; and hardwoods are more preferably used. The wood-derived raw material can be a mixture of softwood and hardwood. A bark can also be used as the wood-derived raw material.

[0073] Examples of hardwoods include beech, Eucalyptus globulus, Eucalyptus grandis, Eucalyptus urograndis, Eucalyptus pellita, Eucalyptus braciana, Acacia mearnsii, and the like. Examples of softwoods include Japanese cedar, Japanese cypress, pine cone, hiba, Japanese hemlock, and the like.

[0074] The wood-derived raw material preferably has a specific gravity of 450 kg/m3 or more and 700 kg/m3 or less, and more preferably 500 kg/m3 or more and 650 kg/m3 or less. When the wood-derived raw material has a specific gravity in the range described above, the production efficiency of acid xylooligosaccharide can be further improved.

[0075] The raw material derived from wood is preferably wood chips obtained by grinding one or more of the previously mentioned woods. When wood chips are used as a plant-derived raw material, the depolymerization of a plant-derived raw material can be efficiently carried out, and the production efficiency of acid xylooligosaccharide can be improved.

Stage I Depolymerization Step

[0076] Step I includes a step of depolymerizing a plant-derived raw material. In the step of depolymerizing a plant-derived raw material, the plant-derived raw material is chemically and/or physically decomposed to produce an acidic xylo-oligosaccharide. Examples of the chemical and/or physical decomposition step include a heat treatment step, an alkali treatment step, an acid treatment step, an enzymatic treatment step, an ionic liquid treatment step, a catalytic treatment step, and the like. . Among these steps, the depolymerization step is preferably a heat treatment step or an enzymatic treatment step; and is more preferably a heat treatment step. The heat treatment step can be a heating and pressurization step.

[0077] The depolymerization step is preferably carried out in non-alkaline conditions (at pH 9 or less, and preferably pH 8 or less).

[0078] The heat treatment step is a step of heating a plant-derived raw material in the presence of a solution. Since the plant-derived raw material is hydrolyzed in a heat treatment step such as this, the heat treatment step is sometimes referred to as a hydrolysis treatment step or a hydrolysis pretreatment step. The solution used in the heat treatment step is preferably water. The ratio (mass ratio) of water to plant-derived raw material is preferably in the range of 1:1 to 1:10. When the ratio of water to plant-derived raw material is established within the range described above, the hydrolysis reaction can be efficiently carried out. The water used in the heat treatment step may be water added separately from the plant-derived raw material; or a portion of the water may be water originally contained in the plant-derived raw material.

[0079] In the heat treatment step, other chemicals can also be added, in addition to the raw material derived from plants and water. Examples of such other chemicals include alkalis, acids, and chelating agents. Additionally, chemicals that directly or indirectly aid in the depolymerization of polysaccharides, such as deposit inhibitors, pitch control agents and ionic liquids can also be added.

[0080] The heat treatment step is a step of heating a plant-derived raw material in the presence of water. The heating temperature (liquid temperature) in this step is preferably 30°C or higher, more preferably 50°C or higher, even more preferably 75°C or higher, even more preferably 90°C or higher, particularly preferably 100°C or higher, and most preferably 120°C or higher. On the other hand, the heating temperature (liquid temperature) is preferably 300°C or lower, more preferably 250°C or lower, and even more preferably 200°C or lower.

[0081] The treatment time in the heat treatment step can be determined, as appropriate, according to the treatment temperature. The treatment time is, for example, preferably 5 minutes or more, more preferably 10 minutes or more, and even more preferably 20 minutes or more. The P factor expressed by the following formula is a product of the heat treatment temperature and the heat treatment time. It is preferable to adjust the P factor within a preferred range.

[0082] In the previous formula, P represents a P factor, T represents an absolute temperature (°C + 273.5), t represents the heat treatment time, and KH1(T)/K1oooc represents the relative rate of bond hydrolysis glycosides.

[0083] In the heat treatment step, the P factor is preferably determined at 2oo or more, more preferably 25o or more, and even more preferably 300 or more. On the other hand, the P factor is preferably 1,000 or less. In the heat treatment step, the P factor is adjusted as appropriate so that the average degree of polymerization and the molecular weight of acidic xylooligosaccharide can be within the desired ranges, while the molecular weight of the pentosan polysulfate obtained can be adjusted.

[0084] In the heat treatment step, the solution containing a plant-derived raw material preferably has a pH of 9 or less, more preferably pH 8 or less, and even more preferably pH 7 or less. That is, the heat treatment step is preferably carried out under non-alkaline conditions. The pH values described above refer to the pH of the solution before heat treatment.

[0085] In the heat treatment step, an acid-derived raw material can be dissociated, and acid hydrolysis can continue at least partially. Examples of raw materials derived from plant acids include organic acids such as acetic acid and formic acid. In this case, the pH of the solution containing a plant-derived raw material is further lowered after acid hydrolysis.

[0086] The method for producing pentosan polysulfate preferably comprises a heat treatment step as the first step. This can improve the production efficiency of acid xylooligosaccharide, and further improves the production efficiency of pentosan polysulfate. When the method includes a heat treatment step as the first step, the number of steps required to produce acidic xylooligosaccharide can be significantly reduced compared to conventional methods. Including a heat treatment under non-alkaline conditions as the first step, the method can efficiently produce acidic xylo-oligosaccharide with suppressed coloring, by virtue of the acidic xylo-oligosaccharide not being replaced with uronic acid.

[0087] The depolymerization step is preferably a heat treatment step; however, it may be a step other than the heat treatment step. For example, when the depolymerization step is an enzymatic treatment step, the depolymerization step includes a step of mixing a plant-derived raw material with an enzyme. Examples of useful enzymes include hemicellulase and the like. Specific examples include commercially available enzyme preparations such as Cellulosin HC100 (brand name, manufactured by HBI Enzymes Inc.), Cellulosin TP25 (brand name, manufactured by HBI Enzymes Inc.), Cellulosin HC (brand name, manufactured by HBI Enzymes Inc.), Cartazyme (brand name, manufactured by Clariant AG), Ecopulp (brand name, manufactured by Rohm Enzima GmbH), Sumizyme (brand name, manufactured by Shin Nihon Chemicals Corporation), Pulpzyme (manufactured by Novo Nordisk), and Multifect 720 (Genencor); and xylanase produced by microorganisms belonging to the genus Trichoderma, genus Thermomyces, genus Aureobasidium, genus Streptomyces, genus Aspergillus, genus Clostridium, genus Bacillus, genus Thermotoga, genus Thermoascus, genus Cardoceram, genus Thermomonospora, or similar.

[0088] In the enzymatic treatment step, an enzyme is added to a solution prepared by mixing a plant-derived raw material with water. The temperature of the solution during this treatment is preferably 10°C or higher and 90°C or lower, and more preferably 30°C or higher and 60°C or lower. The temperature of the solution is preferably a temperature close to the ideal temperature of the enzyme used. The pH of the solution is also preferably adjusted to a range in which enzyme activity is improved. For example, the pH of the solution is preferably adjusted to a pH of 3 or more and a pH of 10 or less.

[0089] When the depolymerization step is an alkaline treatment step or an acid treatment step, the depolymerization step comprises a step of mixing a plant-derived raw material with an alkaline solution or an acid solution. In the alkaline treatment step, sodium hydroxide or potassium hydroxide is preferably added. In the acid treatment step, hydrochloric acid, sulfuric acid, acetic acid, or the like is preferably added. Also in cases like this, heating or pressurization can be carried out, as appropriate.

[0090] When the depolymerization step is at least one selected from an enzymatic treatment step, an alkaline treatment step, and an acid treatment step, the production method may additionally comprise, after the treatment step, a pressing step. , an extraction step, a heating step, a filtration step, a separation step, a purification step, a concentration step, a desalination step, or the like. The method may additionally comprise a molecular weight reduction step carried out after the treatment step. Examples of other steps include the steps described in JP2003-183303A, the contents of which are incorporated herein by reference.

Filtration step

[0091] Step I may additionally comprise a filtration step carried out after the depolymerization step described previously. In the filtration step, the reaction mixture is separated into the solids of the plant-derived raw material, and a solution other than the solids. More specifically, when Step I includes a filtration step performed after the depolymerization step, the reaction product is separated into solids, which are used as a pulp feedstock, and a filtrate. The solids used as a pulp raw material are subjected to a digestion step or the like as a post-step, to provide a cellulose raw material (dissolving pulp).

[0092] The recovered filtrate can be separated into a gas layer and a liquid layer. Since the gas layer contains a large amount of furfurals, furfurals can be isolated by collecting these furfurals from the gas layer. On the other hand, the liquid layer contains a large amount of hemicellulose including acidic xylooligosaccharide and neutral xylooligosaccharide. In the step described below, the acidic xylo-oligosaccharide contained in this liquid layer can be separated and purified.

Separation and Purification Step

[0093] Step I may additionally comprise a separation and purification step carried out after the depolymerization step. When Step I comprises the filtration step described above, a separation and purification step is preferably provided after the filtration step.

[0094] Step I may include a separation and purification step immediately after the depolymerization step. However, step I preferably includes a filtration step carried out after the depolymerization step; and includes a step of separating acidic xylo-oligosaccharide from the filtrate obtained, and purifying the neutral xylo-oligosaccharide. The filtration step may be provided as a part of the separation and purification step; or it may be provided as a step that is independent of the separation and purification step. The separation and purification step is an acid xylooligosaccharide separation and purification step. Since the filtrate obtained in the filtration step contains various saccharides, such as neutral xylo-oligosaccharide, in addition to acidic xylo-oligosaccharide, the separation and purification step is also a step of removing such xylo-oligosaccharides other than xylo- acidic oligosaccharide.

[0095] In the separation and purification step, for example, ion exchange chromatography, affinity chromatography, gel filtration, ion exchange treatment, NF membrane treatment, UF membrane treatment, RO membrane treatment, on activated carbon, or similar methods are preferably used. In the separation and purification step, it is also preferable to carry out the previous methods in combination. In particular, when ion exchange chromatography is performed in separation and purification, acidic xylooligosaccharide can be selectively separated and purified. In ion exchange chromatography, the acidic xylooligosaccharide is adsorbed; In this way, the acidic xylo-oligosaccharide can be mainly obtained from the sugar liquid (filtrate). More specifically, the sugar liquid is first treated with a strong cation exchange resin to remove metal ions from the sugar liquid. Subsequently, using a strong anion exchange resin, sulfate or similar ions are removed from the sugar liquid. The resulting sugar liquid is treated with a weak anion exchange resin to adsorb the acidic xylooligosaccharide onto the resin. An acidic xylooligosaccharide solution with fewer impurities can be obtained by eluting the acidic oligosaccharide adsorbed on the resin with a low salt concentration (e.g., NaCl, CaCl2, KCl or MgCl2).

Concentration Stage

[0096] Step I may additionally comprise a concentration step. The concentration step is preferably provided, for example, after the filtration step, and before the separation and purification step. When step I includes a concentration step like this, the separation and purification step can be more efficiently carried out, thus increasing the pentosan polysulfate production efficiency.

[0097] Examples of the concentration step include a membrane treatment step using an NF membrane, an ultrafiltration membrane, a reverse osmosis membrane, or the like; a concentration step using evaporation etc.; and the like.

[0098] In the concentration step, the solution is preferably concentrated, such that the acidic xylo-oligosaccharide content is 10% or more and 80% or less, and more preferably 20% or more and 60% or less, with based on the total mass of the concentrate.

Dehydration step

[0099] In step I, the acidic xylooligosaccharide can be obtained in the form of an acidic xylooligosaccharide solution; or it can be subjected to a dehydration step and thus obtained in the form of an acid xylo-oligosaccharide concentrate or an acid xylo-oligosaccharide powder. When an acidic xylo-oligosaccharide powder is produced, the production method preferably additionally comprises a powder formation step carried out after the separation and purification step. When a dehydration step is included in the present invention, sulfation in the sulfation step described below can be carried out more efficiently.

[00100] In the powder formation step, the acid xylooligosaccharide solution obtained in the separation and purification step is treated, for example, using a spray dryer, a freeze drying machine, a hot air drying machine , or a water-soluble organic solvent to thus obtain an acidic xylo-oligosaccharide powder.

Stage II Sulfation step

[00101] The acidic xylo-oligosaccharide obtained in step I is sulfated in step II to thus obtain pentosan polysulfate. That is, step II comprises a sulfation step.

[00102] The average degree of polymerization of the acid xylo-oligosaccharide to be subjected to sulfation is preferably adjusted, as appropriate, according to the molecular weight of pentosan polysulfate to be obtained as a final product.

[00103] The average degree of polymerization of acidic xylooligosaccharides can be calculated by dividing the amount of total sugar in the acidic xylooligosaccharide by the amount of reducing sugar. When calculating the amount of total sugar, first, an acidic xylo-oligosaccharide solution is kept at 50°C and centrifuged at 15,000 rpm for 15 minutes. Thereafter, the amount of total sugar in the supernatant is quantified by the phenol sulfuric acid method (“Kangento no Teiryo-Ho (Method of Quantifying Reducing Sugar)”; published by Gakkai Shuppan Center). The calibration curve to be used in quantification is produced using D-xylose (Wako Pure Chemical Industries, Ltd.). The amount of reducing sugar is quantified by the Somogyi-Nelson method (“Kangento no Teiryo-Ho (Method of Quantifying Reducing Sugar)”; published by Gakkai Shuppan Center). The calibration curve to be used in this quantification is also produced using D-xylose (Wako Pure Chemical Industries, Ltd.).

[00104] In the sulfation step, sulfuric acid or a sulfuric acid derivative is added to the acid xylo-oligosaccharide solution to the acid xylo-oligosaccharide sulfate. Examples of sulfuric acid derivatives include pyridine sulfur trioxide complex, chlorosulfonic acid, and the like. In this step, the concentration of the acid xylooligosaccharide solution is preferably 0.1 mass % or more and 20 mass % or less, and sulfuric acid is preferably added to the acid xylooligosaccharide solution having a concentration like this in an amount of 0.1% by mass or more and 50% by mass or less. The acidic xylooligosaccharide solution after addition of sulfuric acid preferably has a pH of 1 or more and 9 or less.

Post-Sulphation Purification Step

[00105] Step II may additionally comprise a post-sulfation purification step carried out after sulfation. When step II includes a post-sulfation purification step like this, a high purity pentosan polysulfate can be obtained.

[00106] In the post-sulfation purification step, for example, centrifugation, membrane filtration, dialysis, water-soluble organic solvent treatment, activated carbon treatment, or similar method is preferably used. Among these, water-soluble organic solvent treatment and activated carbon treatment are preferably used, because pentosan sulfonated polysulfate can be selectively separated and purified.

Powder Formation Step

[00107] In step II, sulfated pentosan polysulfate can be obtained in the form of a pentosan polysulfate solution; or it can be subjected to a powder formation step and thus obtained in the form of a pentosan polysulfate powder. When a pentosan polysulfate powder is produced, step II preferably additionally includes a powder formation step carried out after the post-sulfation purification step.

[00108] In the powder formation step, the pentosan polysulfate solution obtained in the post-sulfation purification step can be treated, for example, using a spray dryer, a freeze drying machine, a hot air drying machine , a water-soluble organic solvent, or the like to thereby obtain a powdered pentosan polysulfate.

[00109] Pentosan polysulfate is obtained by carrying out step II described previously. The pentosan polysulfate thus obtained preferably has a sulfur content of 10% by mass or more to 20% by mass or less, based on the total mass of the pentosan polysulfate. The sulfur content of pentosan polysulfate can be measured by the oxygen bottle combustion method of the General Tests of the Japanese Pharmacopoeia.

Deacetylation Step

[00110] In the production of pentosan polysulfate, deacetylation is preferably carried out. The deacetylation step is preferably carried out at any stage after the depolymerization step. The deacetylation step can reduce the acetyl group content of pentosan polysulfate. Specifically, the deacetylation step is a step of adding a base to a solution containing a substance obtained from a plant-derived raw material, such as acid xylo-oligosaccharide (also referred to herein as a “solution containing acid xylo-oligosaccharide or the like ”), in order to adjust the solution to pH 11 or more. In the deacetylation step, the solution obtained after depolymerization, the filtrate obtained by the filtration step, the solution containing acid xylo-oligosaccharide after the separation and purification step and before the sulfation step, the solution containing acid xylo-oligosaccharide after the sulfation step (pentosan polysulfate), or similar can be adjusted to a pH of 11 or more. Among these solutions, when the solution containing acidic xylooligosaccharide after the separation and purification step and before the sulfation step is adjusted to pH 11 or more, a pentosan polysulfate that has stable quality and a reduced content of acetyl group can be obtained. , and the sites where the acetyl groups were attached can also be sulfated. In this way, the sulfation efficiency, and thus the production efficiency of pentosan polysulfate, can be increased. When the xylo-oligosaccharide-containing solution obtained after the sulfation step (pentosan polysulfate) is adjusted to pH 11 or more, the purification step can be carried out more efficiently. The solution containing acidic xylooligosaccharide or the like is preferably an aqueous solution. The solution containing acidic xylooligosaccharide may also be referred to herein as the acidic xylooligosaccharide solution.

[00111] The pH applied in the deacetylation step is preferably pH 11 to 14, and more preferably pH 12 to 13. The solution to be adjusted for the deacetylation step is preferably maintained at pH 11 or more for 0.5 hour or more , more preferably at pH 11 or more for 1.0 hour or more, even more preferably at pH 11 or more for 2.0 hours or more, and particularly preferably at pH 11 or more for 3.0 hours or more. In particular, when the pH is less than 12, the solution is preferably kept for 1.0 hour or more. Particularly preferred conditions may be, for example, conditions in which the solution is maintained at pH 12 to 13 for 3 hours or more.

[00112] Although the solution is maintained in the pH range described above, the solution is preferably stirred. The temperature applied while the solution is maintained in the pH range described above is not particularly limited, but is preferably room temperature.

[00113] In the deacetylation step, a base can be added to a solution to be subjected to the deacetylation step (a solution containing acidic xylo-oligosaccharide or the like). The base to be added is not particularly limited as long as the desired pH can be achieved. The base is preferably sodium hydroxide.

[00114] The deacetylation step may comprise a pH adjustment step of adjusting, to less than pH 11, the pH of a solution that has a pH of 11 or more, which results from the addition of a base after being maintained at the pH described previously. In the pH adjustment step, the solution may be adjusted to, for example, pH 9 or less, pH 8 or less, pH 7 or less, pH 6 or less, pH 5 or less, or pH 4 or less. Adjustment can be carried out by adding an acid. Examples of useful acids include hydrochloric acid.

[00115] The deacetylation step preferably comprises a desalination step carried out after the pH adjustment step. Desalination can be carried out, for example, using a dialysis membrane or an NF membrane.

[00116] The deacetylation step may additionally comprise a step of powdering the product obtained for subsequent treatment.

Other Steps Molecular Weight Adjustment Step

[00117] The method for producing pentosan polysulfate may additionally comprise a molecular weight adjustment step between step I and step II. When the method for producing pentosan polysulfate includes a deacetylation step, the molecular weight adjustment step can be performed before or after the deacetylation step. In the molecular weight adjustment step, the molecular weight of the acidic xylo-oligosaccharide obtained in step I is adjusted. For example, in the molecular weight adjustment step, the molecular weight of the acidic xylooligosaccharide can be reduced.

[00118] In the molecular weight adjustment step, a pentosan polysulfate having a weight average molecular weight of 1,000 or more and 5,000 or less can be obtained by carrying out, for example, acid treatment, alkaline treatment, enzymatic treatment, membrane treatment, NF, UF membrane treatment, RO membrane treatment, gel filtration treatment, activated carbon treatment, ion exchange treatment, electrodialysis treatment, or the like. It is also possible to use a method of selectively collecting pentosan polysulfate that has a desired weight average molecular weight by carrying out a membrane treatment or the like in the molecular weight adjustment step.

Separation and Purification Step Post-Molecular Weight Adjustment

[00119] The method for producing pentosan polysulfate may additionally comprise a post-molecular weight adjustment separation and purification step carried out after the molecular weight adjustment step. Examples of the post molecular weight adjustment separation and purification step may include gel filtration, ion exchange treatment, NF membrane treatment, UF membrane treatment, RO membrane treatment, electrodialysis treatment, activated carbon treatment , water-soluble organic solvent treatment, chromatographic treatment, and the like. When the production method includes a post-molecular weight adjustment separation and purification step such as this, the acidic xylooligosaccharide that has a desired molecular weight obtained in the molecular weight adjustment step can be selectively collected, and pentosan polysulfate that has a narrow molecular weight distribution can be efficiently obtained.

Examples

[00120] The characteristics of the present invention are described below more specifically with reference to the Production Examples. The materials, amounts used, proportions, treatment contents, treatment procedures, and the like described in the following Production Examples may be appropriately changed as long as such changes do not depart from the spirit of the present invention. Accordingly, the scope of the present invention should not be considered to be limited by the following specific examples.

Production of Acid Xylo-Oligosaccharide (1)

[00121] Fifty parts by mass of water were added to 10 parts by mass of wood chips (hardwood), and a heat treatment was carried out at 165°C for 3 hours. The resulting mixture was then subjected to solid-liquid separation using a Screw Press (manufactured by Shinryo Seisakusho: 250 x 1000 SPH-EN), and the filtrate was recovered. The filtrate was filtered through a 1 μm micron rated bag filter (manufactured by ISP Filters). After 5 parts by mass of activated carbon (PM-SX; manufactured by Mikura Kasei Kabushiki Kaisha) were added to treat the filtrate at 50°C for 2 hours, the resulting mixture, including the activated carbon, was further filtered on a 0.2 μm micron rated ceramic (manufactured by Nihon Pall Co., Ltd.) to recover a clear filtrate. After the clear filtrate was concentrated 20 times with a reverse osmosis membrane (NTR-7450; manufactured by Nitto Denko Corporation) to obtain a concentrated sugar liquid, the concentrated sugar liquid was passed at SV 1.5 through a 4-bed tower type ion exchange resin consisting of a strong cation resin (PK-218; manufactured by Mitsubishi Chemical Corporation), a weak anion resin (WA30; manufactured by Mitsubishi Chemical Corporation), a strong cation resin (PK- 218; manufactured by Mitsubishi Chemical Corporation), and a weak anion resin (WA30; manufactured by Mitsubishi Chemical Corporation). The acidic xylooligosaccharide was thus adsorbed on the weak anionic resin of the second and fourth towers. A 50 mM aqueous sodium chloride solution was then passed through the second and fourth towers at SV 1.5 to recover an acidic xylooligosaccharide solution. Sodium hydroxide was added to the obtained acid xylooligosaccharide solution to reach a pH of 13, and the resulting mixture was stirred at room temperature for 3 hours for deacetylation. After hydrochloric acid was added to the resulting solution to achieve a pH of less than 5 and desalination was performed using a dialysis membrane (Spectra/Por 7, EC membrane, MWCO 100-500; manufactured by Spectrum), the resulting mixture was sprayed using a freeze drying machine (manufactured by EYELA).

Production of Neutral Xylooligosaccharide

[00122] Fifty parts by mass of water were added to 10 parts by mass of wood chips (hardwood), and a heat treatment was carried out at 165°C for 3 hours. The resulting mixture was then subjected to solid-liquid separation using a Screw Press (manufactured by Shinryo Seisakusho: 250 x 1000 SPH-EN), and the filtrate was recovered. The filtrate was further filtered on a 1 μm micron rated bag filter (manufactured by ISP Filters). After 5 parts by mass of activated carbon (PM-SX; manufactured by Mikura Kasei Kabushiki Kaisha) were added to treat the filtrate at 50°C for 2 hours, the resulting mixture, including the activated carbon, was further filtered on a 0.2 μm micron rated ceramic (manufactured by Nihon Pall Co., Ltd.) to recover a clear filtrate. After the clear filtrate was concentrated 20 times with a reverse osmosis membrane (NTR-7450; manufactured by Nitto Denko Corporation) to obtain a concentrated sugar liquid, the concentrated sugar liquid was passed at SV 1.5 through a 4-bed tower type ion exchange resin consisting of a strong cation resin (PK-218; manufactured by Mitsubishi Chemical Corporation), a weak anion resin (WA30; manufactured by Mitsubishi Chemical Corporation), a strong cation resin (PK- 218; manufactured by Mitsubishi Chemical Corporation), and a weak anionic resin (WA30; manufactured by Mitsubishi Chemical Corporation) to thus recover a neutral xylo-oligosaccharide solution. Sodium hydroxide was added to the obtained neutral xylooligosaccharide solution to reach a pH of 13, and the resulting mixture was stirred at room temperature for 3 hours for deacetylation. After hydrochloric acid was added to the solution obtained to reach a pH less than 5, and the salt obtained was removed using a dialysis membrane (Spectra/Por 7, EC membrane, MWCO 100-500; manufactured by Spectrum), the mixture resulting was pulverized using a freeze drying machine (manufactured by EYELA).

Production of Pentosan Sodium Polysulfate Comparative example 1

[00123] 25 mL of N,N-dimethylformamide, 12.4 g of sulfur trioxide and pyridine complex, and 1.5 g of neutral xylo-oligosaccharide powder produced by the method described previously were placed in a separation flask. 100 mL, and a reaction continued at 40°C for 3 hours. After cooling, the obtained reaction mixture was added dropwise to 500 mL of ethanol. The generated precipitate was collected by filtration, and 30 mL of water was added to dissolve the precipitate in it. A sodium hydroxide solution was added to the solution to reach a pH of 10. The resulting solution was added dropwise to 500 mL of ethanol, and the precipitate obtained was then collected by filtration. Thereafter, 30 mL of water was added to dissolve the precipitate in it; and activated carbon was added to the solution and stirred, followed by filtration. The filtrate was concentrated using an evaporator, and pulverized using a freeze drying machine (manufactured by EYELA).

Comparative Example 2

[00124] Sodium pentosan polysulfate was obtained in the same way as in Comparative Example 1, except that a mixture of 1.125 g of powdered neutral xylo-oligosaccharide and 0.375 g of acidic xylo-oligosaccharide (1) was used to instead of 1.5 g of the powdered neutral xylo-oligosaccharide of Comparative Example 1.

Example 1

[00125] Sodium pentosan polysulfate was obtained in the same way as in Comparative Example 1, except that a mixture of 0.375 g of powdered neutral xylo-oligosaccharide and 1.125 g of acidic xylo-oligosaccharide (1) was used to instead of 1.5 g of the powdered neutral xylo-oligosaccharide of Comparative Example 1.

Example 2

[00126] Sodium pentosan polysulfate was obtained in the same way as in Comparative Example 1, except for the fact that 1.5 g of acidic xylo-oligosaccharide (1) was used instead of 1.5 g of neutral xylo-oligosaccharide powder from Comparative Example 1.

Physical Property Values

[00127] The uronic acid content, average molecular weight, and sulfur content of pentosan polysulfates of Examples 1 and 2 and Comparative Examples 1 and 2 were measured as follows.

Uronic acid content

[00128] About 10 mg of sodium pentosan polysulfate obtained in each of the Examples and Comparative Examples were weighed and dissolved in distilled water to prepare the volume exactly 25 mL. 1 mL of each solution was placed in a test tube. While the solution was cooled in ice water, 5 mL of a 0.025 M sodium tetraborate solution in sulfuric acid was added and mixed, and the resulting mixture was heated in a water bath for 10 minutes. Immediately after heating, the resulting mixture was cooled on ice, and 0.2 mL of a carbazole reagent was added and mixed. The resulting mixture was heated in a water bath for 15 minutes, and then cooled naturally to obtain a sample solution. Separately, standard stock solutions of glucuronic acid at a concentration of 10 to 100 μg/mL were prepared and subjected to the same procedure as previously to obtain standard solutions. 1 mL of distilled water was also subjected to the same procedure, and the resulting liquid was used as a control. The absorbance at a wavelength of 530 nm was measured. Calibration curves were prepared from the absorbance of the standard solutions, and the amount of glucuronic acid (g) in the Examples and Comparative Examples was determined. The uronic acid content (% by mass) was calculated according to the following formula. When the quantitative value was negative, it was considered as 0%. Uronic acid content (% by mass) = Amount of glucuronic acid (μg)/(Amount of pentosan sodium polysulfate weighed x 1/25)/10

Sulphur content

[00129] The sulfur content was measured by the oxygen bottle combustion method described in the Japanese Pharmacopoeia.

Average molecular weight

[00130] The weight average molecular weight (Mw) of the pentosan polysulfate of the present invention can be determined by GPC (gel permeation chromatography). A YMC-Pack Diol-300 and YMC-Pack Diol-60 (both manufactured by YMC) connected to each other can be used as a GPC column. GPC was carried out, for example, under the following conditions.

[00131] Eluent: 25 mM potassium dihydrogen phosphate / 25 mM dipotassium hydrogen phosphate / 50 mM potassium chloride Flow rate: 0.7 mL/min Measuring temperature: 40°C Detector: refractive index detector

Inhibitory Activity to Inhibit the Binding Between FGF-2 and Heparan Sulfate

[00132] 10 mg of pentosan polysulfate from each of the Examples and Comparative Examples were dissolved in 10 mL of saline. The resulting solutions were mixed with biotinylated heparan sulfate and stirred at 37°C for 15 minutes. As a blank, a sample obtained by mixing only saline with biotinylated heparan sulfate was also prepared. These solutions were added to a plate immobilized with FGF-2, and the plate was shaken at 37°C for 15 minutes. After the solutions were removed and the plate wells were washed with 0.1% Tween 20/PBS three times, an avidin-HRP solution was added, and the plate was shaken at 37°C for 15 minutes. After the solution was removed and the plate wells were washed with 0.1% Tween 20/PBS three times, a substrate for HRP was added to allow color to appear at room temperature for about 5 minutes. After a reaction stop solution was added and stirred, the absorbance at a wavelength of 450 nm (A450 nm) was measured. The inhibition rate was calculated by the following formula. Inhibition rate (%) = (A450 nm (white) - A450 nm (pentosan polysulfate)) / A450 nm (white)

[00133] The biotinylated heparan sulfate, FGF-2 immobilized plate, avidin-HRP solution, and HRP substrate used were those included in a Heparan Degradation Enzyme Assay Kit (Takara Bio Inc.). Tween 20, PBS, and the reaction stop solution used were those included in a Sulfuric Acid Free ELISA Wash and Stop Solution (Takara Bio Inc.).

[00134] Table 1 and figure 1 show the results. Table 1

[00135] The results in table 1 and figure 1 show that pentosan sodium polysulfate that has a uronic acid content of 7.0% by mass to 15.0% by mass presents a high FGF inhibition rate.

Production of Acid Xylooligosaccharide (2)

[00136] Fifty parts by mass of water were added to 10 parts by mass of wood chips (hardwood), and a heat treatment was carried out at 165°C for 3 hours. The resulting mixture was then subjected to solid-liquid separation using a Screw Press (manufactured by Shinryo Seisakusho: 250 x 1000 SPH-EN), and the filtrate was recovered. The filtrate was further filtered on a 1 μm micron rated bag filter (manufactured by ISP Filters). After 5 parts by mass of activated carbon (PM-SX; manufactured by Mikura Kasei Kabushiki Kaisha) were added to treat the filtrate at 50°C for 2 hours, the resulting mixture, including the activated carbon, was further filtered on a 0.2 μm micron rated ceramic (manufactured by Nihon Pall Co., Ltd.) to recover a clear filtrate. After the clear filtrate was concentrated 20 times with a reverse osmosis membrane (NTR-7450; manufactured by Nitto Denko Corporation) to obtain a concentrated sugar liquid, the concentrated sugar liquid was passed at SV 1.5 through a 4-bed tower type ion exchange resin consisting of a strong cation resin (PK-218; manufactured by Mitsubishi Chemical Corporation), a weak anion resin (WA30; manufactured by Mitsubishi Chemical Corporation), a strong cation resin (PK- 218; manufactured by Mitsubishi Chemical Corporation), and a weak anion resin (WA30; manufactured by Mitsubishi Chemical Corporation). The acidic xylooligosaccharide was thus adsorbed on the weak anionic resin of the second and fourth towers. A 50 mM aqueous sodium chloride solution was then passed through the second and fourth towers at SV 1.5 to recover an acidic xylooligosaccharide solution. The obtained acid xylo-oligosaccharide solution was pulverized using a freeze-drying machine (manufactured by EYELA).

Production of Acid Xylooligosaccharide (3)

[00137] An acidic xylo-oligosaccharide (3) was obtained in the same way as in the production of the acidic xylo-oligosaccharide (1), except for the fact that deacetylation was carried out at pH 11 for 1 hour.

Production of Acid Xylooligosaccharide (4)

[00138] An acidic xylo-oligosaccharide (4) was obtained in the same way as in the production of the acidic xylo-oligosaccharide (1), except for the fact that deacetylation was carried out at pH 11 for 2 hours.

Production of Acid Xylooligosaccharide (5)

[00139] An acidic xylo-oligosaccharide (5) was obtained in the same way as in the production of the acidic xylo-oligosaccharide (1), except for the fact that deacetylation was carried out at pH 12 for 1 hour.

Production of Pentosan Sodium Polysulfate 2 Comparative Example 3

[00140] Pentosan sodium polysulfate was obtained in the same way as in Comparative Example 1, except for the fact that 1.5 g of acidic xylo-oligosaccharide powder (2) was used instead of 1.5 g of xylo - powdered neutral oligosaccharide from Comparative Example 1.

Examples 3 to 5

[00141] The sodium pentosan polysulfate of each of Examples 3 to 5 was obtained in the same way as in Comparative Example 1, except that 1.5 g of acidic xylooligosaccharides (3) to (5) was used at the same time. instead of 1.5 g of the powdered neutral xylo-oligosaccharide of Comparative Example 1.

Physical Properties and Inhibitory Activity to Inhibit the Binding Between FGF-2 and Heparan Sulfate

[00142] The uronic acid content and the average molecular weight of the pentosan polysulfates of Examples 3 to 5 and Comparative Example 3 were determined in the same way as in Example 1. The acetyl group content of the pentosan polysulfates of Examples 2 to 5 and Example Comparative 3 was determined as follows.

[00143] 35 mg of sodium 3-(trimethylsilyl)propionate-2,2,3,3-d4 (produced by Isotec Corporation) was dissolved in heavy water (produced by Kanto Kagaku). Using a 25 mL measuring bottle, the solution was diluted to prepare an internal standard solution. The sodium pentosan polysulfate obtained in each of the Examples and Comparative Examples was weighed (30 mg) and dissolved in 1 mL of the internal standard solution to prepare a solution for NMR measurement. The obtained solution was transferred to an NMR sample tube (produced by Kanto Kagaku), and HNMR measurement was performed using FT-NMR (JNM-LA400; produced by JEOL Ltd.). The acetyl group content was calculated from the integral ratio of the peak for the trimethylsilyl group of the internal standard substance and the peak for the acetyl group of sodium pentosan polysulfate.

[00144] The uronic acid content, average molecular weight and inhibitory activity to inhibit the binding between FGF-2 and pentosan heparan sulfate polysulfates of Examples 2 to 5 and Comparative Example 3 were determined in the same way as in Example 1 , table 2 and figure 2 show the results. Table 2

Storage Stability

[00145] Pentosan sodium polysulfate from each of Examples 2 and 4 and Comparative Example 3 was dissolved in purified water at a concentration of 100 mg/mL, and the resulting solutions were sealed in screw bottles. The properties of each solution after storage at 40°C for 1 week and 2 weeks were visually confirmed. Table 3

[00146] The results in table 2 and figure 2 show that any sodium pentosan polysulfate that has an acetyl group content of less than 2.0% had an FGF inhibition rate of more than 90%, thus presenting a high rate of inhibition. Table 3 shows that when aqueous solutions of sodium pentosan polysulfate that have a low acetyl group content are stored at room temperature or higher, coloration hardly occurs and the solutions are highly stable.

pH Buffering Action of Pentosan Polysulfate

[00147] 100 mg of pentosan polysulfate obtained in each of Examples 1 and 2 and Comparative Examples 1 and 2 were dissolved in water to prepare the total volume of exactly 100 mL. This solution was adjusted to pH 10 using 0.01N aqueous sodium hydroxide solution (produced by Kanto Kagaku) with an automatic titrator (produced by DKK Toa Corporation). The titration was then performed using an aqueous solution of 0.01N hydrochloric acid (produced by Kanto Kagaku) with the automatic titrator. The amount of 0.01N aqueous hydrochloric acid solution required to adjust the pH of the pentosan polysulfate solution from pH 6 to pH 4 was calculated.

[00148] Figure 3 shows the results.

[00149] The results in figure 3 clearly showed that the pentosan polysulfates of the Examples have strong buffering action to maintain the pH in the range of pH 4 to pH 6.

Claims (14)

1. Pentosan polysulfate, characterized by the fact that it has a uronic acid content of 7.0% by mass to 15.0% by mass, and an acetyl group content of 0% by mass to 1.62% by mass; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 2. Pentosan polysulfate according to claim 1, characterized in that the pentosan polysulfate has a uronic acid content of 7.5% by mass to 13.0% by mass; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 3. Pentosan polysulfate according to claim 1 or 2, characterized in that the pentosan polysulfate has a weight average molecular weight of 5,000 or less; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 4. Pentosan polysulfate according to claim 3, characterized in that the pentosan polysulfate has an acetyl group content of 0 to 0.3% by mass; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 5. Pentosan polysulfate according to any one of claims 1 to 4, characterized by the fact that the pentosan polysulfate has a structure represented by Formula II: wherein R, each independently, represents a hydrogen atom, -COCH3, or -SO3X1, and at least one R in the molecule is -SO3X1, wherein X1 represents a hydrogen atom or a monovalent or divalent metal; X represents a hydrogen atom or a monovalent or divalent metal; and n1 and n2, each independently, represent an integer of 0 or more and 30 or less, and at least one of n1 and n2 is an integer of 1 or more; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 6. Pentosan polysulfate according to claim 5, characterized in that each R independently represents a hydrogen atom or -SO3X1; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 7. Pentosan polysulfate according to claim 5, characterized in that X1 is sodium; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 8. Medication, characterized by the fact that it comprises pentosan polysulfate as an active ingredient as defined in any one of claims 1 to 7; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 9. Use of pentosan polysulfate as defined in any one of claims 1 to 7, a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or the pharmaceutically acceptable salt thereof, characterized in that it is for the manufacture of a medicament for the prevention and/or treatment of a disease caused by an abnormal increase in FGF-2 function. 10. Use according to claim 9, characterized in that the disease caused by abnormal increase in FGF-2 function is cancer, autoimmune disease, allergic disease, inflammatory disease, cardiac dysplasia, vascular dysplasia or skeletal dysplasia. 11. Use according to claim 9, characterized by the fact that the disease caused by the abnormal increase in FGF-2 function is cystitis or arthritis. 12. Medication according to claim 8, characterized by the fact that it is an injectable formulation. 13. pH buffering agent, characterized by the fact that it comprises pentosan polysulfate as defined in any one of claims 1 to 7; a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof. 14. Use of pentosan polysulfate as defined in any one of claims 1 to 7, a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable solvate of pentosan polysulfate or pharmaceutically acceptable salt thereof, characterized in that it is as a pH buffering agent.