Folate for medical use
Patent Information
- Authority / Receiving Office
- JP · JP
- Patent Type
- Patents
- Current Assignee / Owner
- APROFUL CO LTD
- Filing Date
- 2023-05-24
- Publication Date
- 2026-08-05
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Figure 0007901046000001 
Figure 0007901046000002 
Figure 0007901046000003
Abstract
Description
[Technical Field]
[0001] This invention relates to folate, its preparation, and compositions containing the same. [Background technology]
[0002] Depression, along with other mental health disorders such as dementia, autism, ADHD, and Alzheimer's disease, as well as non-communicable diseases (NCDs) such as type 2 diabetes and vascular disease, and cancer, are increasing concerns for patients and healthcare systems, especially considering the elderly population. While there are various reasons for these diverse conditions, a common risk factor has been identified: suboptimal folate levels throughout the body or in specific tissues.
[0003] For example, it is well known that B vitamins are involved in many metabolic processes in the body, such as converting carbohydrates into glucose, which is metabolized for energy production. These vitamins are also essential for the breakdown of fats and proteins. Furthermore, they play an important role in maintaining muscle tone along the digestive tract and promoting the health of the nervous system and organs such as the eyes, skin, hair, and liver.
[0004] Furthermore, folic acid is known to be essential for the generation and maintenance of new cells. It is especially important during periods of rapid cell division and growth, such as infancy and pregnancy. Folic acid is required for DNA replication. Therefore, folic acid deficiency most clinically affects the bone marrow and other areas of rapid cell turnover, interfering with DNA synthesis and cell division. Since RNA and protein synthesis are not affected, megaloblasts are produced, leading to macrocytic anemia, such as megaloblastic anemia. This can be seen in celiac disease, nutritional anemia, or during pregnancy, infancy, or childhood. Therefore, both adults, especially the elderly, and children need folic acid to produce normal red blood cells and prevent anemia. Folic acid also helps prevent DNA changes that can lead to cancer.
[0005] Folic acid derivatives, such as various tetrahydrofolate derivatives, can also be used as drugs or as basic substances for preparing other derivatives. However, tetrahydrofolate and its derivatives are also known to be extremely unstable, particularly due to their susceptibility to oxidation. Specifically, 5-formyltetrahydrofolate (folic acid, leucovorin) is important as a drug component in cancer research and in the treatment of folate deficiency anemia associated with antibiotic therapy during pregnancy, as a therapy adjunct to methotrexate and 5-fluorouracil. Calcium salts can be mentioned as the most relatively stable derivatives of folate and reduced folate. U.S. Patents 5,817,659 and 6,441,168 disclose calcium salts of 5-methyl-(6R,S)-, (6S)-, or (6R)-tetrahydrofolate, preferably having at least 1 equivalent of crystalline water per equivalent of the acid. 5-methyltetrahydrofolate is the only commercially available folate derivative that can directly cross the blood-brain barrier without further metabolism. Natural 5-methyltetrahydrofolate exists only in the S form; the R form is biochemically inactive and is thought to be excreted via the kidneys. In addition, several compositions for human and animal intake containing either folate and / or reduced folate have been reported, in various forms, along with vitamins, arginine, lysine, thiamine, and / or other active ingredients, either as nutritional supplements or for the treatment and prevention of various diseases, such as neurological, pathophysiological cardiovascular diseases, arthritis, or inflammatory conditions.
[0006] Various folate salts are known. Generally, these salts contain folic acid and inorganic cations such as calcium and magnesium, or organic cations such as glucosamine or galactosamine. These alkaline earth metal cations are inert unless the cation itself exhibits any pharmacological effect in humans. The poor solubility of these salts in aqueous solutions has been widely reported. International Publication No. 2009 / 103334 describes 5-methyltetrahydrofolate glucosamine salt (5-MTHF-glucosamine), which has good water solubility. Furthermore, the solubility of folate salts in nonpolar solvents is also very limited, similar to that in aqueous solutions. For example, 5-MTHF-glucosamine was described as a cream-colored to light brown powder that is highly soluble in water (25°C), soluble in dilute acids or alkalis, and insoluble in organic solvents (EFSA Journal 2013;11(10):3358). Aqueous compositions of folic acid with improved solubility and stability are disclosed, for example, in U.S. Patent Nos. 9,301,922 and 9,642,853. Furthermore, many folic acid compositions have been described that include folic acid and further compounds such as vitamins, lysine, thiamine, and other active ingredients. However, combinations of further active compounds and stable folate salts with good solubility in water and nonpolar solvents would provide even more versatile pharmaceutical compositions. [Overview of the project]
[0007] The objective of the present invention is to provide a folate salt that combines further active compounds and exhibits good stability, high solubility in nonpolar solvents, and good water solubility.
[0008] This objective is achieved by the folate according to the present invention as defined in claim 1. More preferred embodiments are provided for in the dependent claims.
[0009] The amorphous folate according to the present invention consists of a tetrahydrofolate anion and an organic cation, and this anion is selected from the group consisting of 5-formyl-(6S)-tetrahydrofolate, 10-formyl-(6R)-tetrahydrofolate, 5-methyl-(6S)-tetrahydrofolate, (6S)-tetrahydrofolate, 5,10-diformyl-(6S)-tetrahydrofolate, 5-methyl-10-formyl-(6S)-tetrahydrofolate, 5,10-methylene-(6R)-tetrahydrofolate and its oxidation derivatives JK12A and Mefox. Further, the cation is an organic compound selected from the group consisting of arginine, choline, acetylcholine, 1,1-dimethyl-biguanidine, phenylethylbiguanidine, betaine-methyl ester and dimethylaminoethanol. The folate has a high solubility in a nonpolar solvent, and the nonpolar solvent has a relative polarity of 0.82 compared to water.
[0010] Higher than the solubility of a specific folate at 5 mass percent (m / m) in a specific nonpolar solvent is regarded as a high solubility in the nonpolar solvent. The solubility was measured at 20 °C.
[0011] The structure of Mefox is shown below.
Chemical formula
[0012] The structure of JK12A is shown below.
Chemical formula
[0013] Compounds such as Mefox and JK12A are both oxidation derivatives of 5-methyl-(6S)-tetrahydrofolate.
[0014] In a preferred embodiment, the amorphous folate consists of a tetrahydrofolate anion and an organic cation. The anion is selected from the group consisting of 5-formyl-(6S)-tetrahydrofolate, 10-formyl-(6R)-tetrahydrofolate, 5-methyl-(6S)-tetrahydrofolate, (6S)-tetrahydrofolate, 5,10-methylene-(6R)-tetrahydrofolate and its oxidation derivatives JK12A and Mefox. The cation is selected from the group consisting of di-arginine, di-choline, di-acetylcholine, di-(1,1-dimethyl-biguanidine), di-(phenyl-ethyl-biguanidine), di-betaine-methyl ester and di-dimethyl-amino-ethanol.
[0015] In another embodiment, the amorphous folate consists of a tetrahydrofolate anion and an organic cation, the anion is selected from the group consisting of 5-formyl-(6S)-tetrahydrofolate, 10-formyl-(6R)-tetrahydrofolate, 5-methyl-(6S)-tetrahydrofolate, (6S)-tetrahydrofolate, 5,10-methylene-(6R)-tetrahydrofolate and its oxidation derivatives JK12A and Mefox, and the cation is selected from the group consisting of mono-arginine, mono-choline, mono-acetylcholine, mono-(1,1-dimethyl-biguanidine), mono-(phenyl-ethyl-biguanidine)), mono-betaine-methyl ester and mono-dimethyl-amino-ethanol.
[0016] In a further embodiment, the amorphous tetrahydrofolate consists of 5-formyl-(6S)-tetrahydrofolate as the tetrahydrofolate anion and mono-arginine as the organic cation. In this case, the 1 1H-NMR shifts in D2O are as follows.
Table 1
[0017] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of mono-(1,1-dimethyl-biguanidine). In this case, the D2O 1 The H-NMR shifts are as follows: [Table 2]
[0018] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of mono-(phenylethyl-biguanidine). In this case, in D2O 1 The H-NMR shifts are as follows: [Table 3]
[0019] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of monocholine. In this case, in D2O 1 The H-NMR shifts are as follows: [Table 4]
[0020] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of mono-(1,1-dimethyl-biguanidine). In this case, the D2O 1 The H-NMR shifts are as follows: [Table 5]
[0021] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of mono-(phenylethyl-biguanidine). In this case, the 1 1H-NMR shifts in D2O are as follows. [Table 6]
[0022] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of mono-choline. In this case, the 1 1H-NMR shifts in D2O are as follows. [Table 7]
[0023] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of mono-2-dimethylaminoethanol. In this case, the 1 1H-NMR shifts in D2O are as follows. [Table 8]
[0024] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of mono-arginine. In this case, the 1 1H-NMR shifts in D2O are as follows. [Table 9]
[0025] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of mono-acetylcholine. In this case, in D2O 1 The H-NMR shifts are as follows: [Table 10]
[0026] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of mono-acetylcholine. In this case, in D2O 1 The H-NMR shifts are as follows: [Table 11]
[0027] In a further embodiment, the amorphous tetrahydrofolate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of di-arginine. In this case, the D2O 1 The H-NMR shifts are as follows: [Table 12]
[0028] In a further embodiment, the amorphous folate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of dicholine. In this case, the D2O 1 The H-NMR shifts are as follows: [Table 13]
[0029] In another embodiment, the amorphous folate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of di-(1,1-dimethyl-biguanidine). In this case, the D2O 1 The H-NMR shifts are as follows: [Table 14]
[0030] In a further embodiment, the amorphous folate consists of a tetrahydrofolate anion of 5-formyl-(6S)-tetrahydrofolate and an organic cation of di-(phenylethyl-biguanidine). In this case, in D2O 1 The H-NMR shifts are as follows: [Table 15]
[0031] In another embodiment, the amorphous folate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of diarginine. In this case, the D2O 1 The H-NMR shifts are as follows: [Table 16]
[0032] In another embodiment, the amorphous folate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of di-(1,1-dimethyl-biguanidine). In this case, the D2O 1 The H-NMR shifts are as follows: [Table 17]
[0033] In a further embodiment, the amorphous folate consists of a tetrahydrofolate anion of 5-methyl-(6S)-tetrahydrofolate and an organic cation of di-(phenylethyl-biguanidine). In this case, in D2O 1 The H-NMR shifts are as follows: [Table 18]
[0034] In a more preferred embodiment, the pharmaceutical composition comprises at least one folate according to the present invention as the main active compound. The composition further comprises at least pharmaceutically acceptable excipients. The composition may, for example, contain buffering compounds. Preferred buffering compounds are trometamol and HEPES. Furthermore, antioxidant compounds may be present in the composition. Preferred antioxidant compounds are thioglycerol, dithiothreitol (DTT), and cysteine.
[0035] Furthermore, at least one folate according to the present invention can be used for the preparation of drugs, food additives, or nutritional supplements, and for the prevention and / or treatment of any deficiency or disorder that is positively affected by the administration of folate. Many disease symptoms are positively affected by compositions containing folate. Such diseases include, for example, pathophysiological, neurological, and inflammatory diseases.
[0036] Furthermore, the method for preparing amorphous folate according to the present invention comprises the step of adding oxalic acid or, alternatively, a fluoride salt to an aqueous composition of an alkaline earth metal salt of folate, wherein the tetrahydrofolate consists of a folate anion and an organic cation. example Example 1 Preparation of 5-formyl-(6S)-tetrahydrofolate di-L-arginine salt Under argon light, 10.0 g (16.62 mmol) of calcium levofolate pentahydrate was dissolved in 230 ml of water at 70°C. Next, 5.79 g (33.24 mmol) of L-arginine was added, followed by 2.09 g (16.62 mmol) of oxalic acid dihydrate. After cooling to room temperature (RT), the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 6) was evaporated to dryness. The crude product was then filtered by warm immersion in methanol, and dried under vacuum at 60°C to obtain 13.63 g of the title compound. Analytical data: [Table 19] Optical rotation:α 20D +1.75°(c=1H2O) Example 2 Preparation of 5-formyl-(6S)-tetrahydrofolate dimethformin salt Under argon, 1.86 g (3.09 mmol) of calcium levofolate pentahydrate was dissolved in 43 ml of water at 70°C. Next, 0.80 g (6.18 mmol) of metformin free base was added, followed by 0.39 g (3.09 mmol) of oxalic acid dihydrate. After cooling to room temperature, the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 6) was evaporated to dryness. The crude product was then filtered by warm immersion in ethanol, and dried under vacuum at 60°C to obtain 2.13 g of the title compound. Analytical data: [Table 20] Optical rotation:α 20D - 2.84°(c=1H2O) Example 3 Preparation of 5-formyl-(6S)-tetrahydrofolate diphenformine salt Under argon, 1.21 g (2.02 mmol) of calcium levofolate pentahydrate was dissolved in 43 ml of water at 70°C. Next, 0.83 g (4.04 mmol) of phenformin free base was added, followed by 0.25 g (2.02 mmol) of oxalic acid dihydrate. After cooling to room temperature, the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 5.5) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile, and dried under vacuum at 60°C to obtain 1.80 g of the title compound. Analytical data: [Table 21] Optical rotation:α 20D +1.39°(c=1H2O) Example 4 Preparation of 5-formyl-(6S)-tetrahydrofolate dicholine salt Under argon, 1.92 g (3.19 mmol) of calcium levofolate pentahydrate was dissolved in 40 ml of water at 70°C. Next, an aqueous solution of 0.79 g (6.38 mmol) of filtered choline fluoride (obtained from choline chloride and silver fluoride) was added. After cooling to 0°C, the precipitated calcium fluoride was filtered off using a high-flow pad, and the resulting clear solution (pH 6.5) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile, and dried under vacuum at 60°C to obtain 1.89 g of the title compound. Analytical data: [Table 22] Optical rotation:α 20D +1.14° (c=1H2O) Example 5 Preparation of 5-methyl-(6S)-tetrahydrofolate di-L-arginine salt Under argon, 10.0 g (17.67 mmol) of levome calcium folate 3.8-hydrate was suspended in 100 ml of water at 70°C. Next, 2.23 g (17.67 mmol) of oxalic acid dihydrate was added, followed by 6.15 g (35.33 mmol) of L-arginine. After cooling to 0°C, the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 6) was evaporated to dryness. The crude product was then filtered by warm immersion in methanol, and dried under vacuum at 60°C to obtain 13.98 g of the title compound. Analytical data: [Table 23] Optical rotation:α 20D +30.0° (c=1H2O) Example 6 Preparation of 5-methyl-(6S)-tetrahydrofolate dimethformin salt Under argon, 1.49 g (2.63 mmol) of levome calcium folate 3.8-hydrate was suspended in 38 ml of water at 70°C. Next, 0.332 g (2.63 mmol) of oxalic acid dihydrate dissolved in water was added, followed by 0.68 g (5.26 mmol) of metformin free base dissolved in water. After cooling to 0°C, the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 6) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile, and dried under vacuum at 60°C to obtain 1.89 g of the title compound. Analytical data: [Table 24] Optical rotation:α 20D +17.95° (c=1H2O) Example 7 Preparation of 5-methyl-(6S)-tetrahydrofolate diphenformin salt Under argon, 1.64 g (2.90 mmol) of levome calcium folate 3.8-hydrate was suspended in 35 ml of water at 70°C. Next, 0.366 g (2.90 mmol) of oxalic acid dihydrate dissolved in water was added, followed by 0.68 g (5.26 mmol) of phenformin free base dissolved in water. After cooling to 0°C, the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 5.5) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile, and dried under vacuum at 60°C to obtain 2.54 g of the title compound. Analytical data: [Table 25] Optical rotation:α 20D +16.06° (c=1H2O) Example 8 Preparation of 5-formyl-(6S)-tetrahydrofolate mono-L-arginine salt Under argon light, 0.258 g (0.429 mmol) of calcium levofolate pentahydrate was dissolved in 6 ml of 70°C water. Next, 0.0747 g (0.429 mmol) of L-arginine was added, followed by 0.0541 g (0.429 mmol) of oxalic acid dihydrate dissolved in water. After cooling to room temperature, the precipitated calcium oxalate was filtered off using a high-flow pad, and the resulting clear solution (pH 4) was evaporated to dryness. The crude product was then filtered by warm immersion in methanol, dried under vacuum at 60°C, and 0.274 g of the title compound was obtained. Analytical data: [Table 26] Optical rotation:α 20D +0.62° (c=1H2O) Example 9 Preparation of 5-formyl-(6S)-tetrahydrofolate monomethformin salt Under argon, 100.0 mg (0.2112 mmol) of levofolinic acid and 28.1 mg (0.2175 mmol) of metformin free base were suspended in 2 ml of water and 4 ml of methanol. The mixture was stirred for 15 minutes, and after sonication, the resulting solution (pH 5) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile and dried under vacuum at 60°C to obtain 122 mg of the title compound. Analytical data: [Table 27] Optical rotation:α 20D -15.3° (c=1H2O) Example 10 Preparation of 5-formyl-(6S)-tetrahydrofolate monophenformine salt Under argon, 100.1 mg (0.2114 mmol) of levofolinic acid and 44.7 mg (0.2178 mmol) of phenformin free base were suspended in 4 ml of water and 4 ml of methanol. After stirring and sonication for 10 minutes, the resulting solution (pH 5) was evaporated to dryness. The crude product was then filtered by warm immersion with acetonitrile and dried under vacuum at 60°C to obtain 137 mg of the title compound. Analytical data: [Table 28] Optical rotation:α 20D +13.50° (c=1H2O) Example 11 Preparation of 5-formyl-(6S)-tetrahydrofolate monocholine salt Under argon, 100.0 mg (0.2112 mmol) of levofolinic acid and 62.5 µl (0.2218 mmol) of a 45% methanol solution of choline hydroxide were suspended in 2 ml of water and 4 ml of methanol. After stirring for 5 minutes, sonication was performed, and the mixture was heated under reflux. The resulting slightly turbid solution (pH 5) was filtered using a syringe filter while warming, and evaporated to dryness. Next, the crude product was filtered by warm immersion in acetonitrile, dried under vacuum at 60°C, and 118 mg of the title compound was obtained. Analytical data: [Table 29] Optical rotation:α 20D -16.3° (c=1H2O) Example 12 Preparation of 5-methyl-(6S)-tetrahydrofolate monomethformin salt Under argon, 72.7 mg (0.1582 mmol) of levomefolic acid and 21.5 mg (0.1661 mmol) of metformin free base were suspended in 10 ml of water and 20 ml of methanol. The mixture was stirred, sonicated, refluxed, and the resulting solution was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile and dried under vacuum at 50°C to obtain 94.6 mg of the title compound. Analytical data: [Table 30] Example 13 Preparation of 5-methyl-(6S)-tetrahydrofolate monophenformine salt Under argon, 90.4 mg (0.1967 mmol) of levomefolic acid and 42.4 mg (0.2066 mmol) of phenformin free base were suspended in 5 ml of water and 10 ml of methanol. The mixture was stirred, sonicated, and refluxed for a period of time. The resulting solution was then evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile and dried under vacuum at 50°C to obtain 133 mg of the title compound. Analytical data: [Table 31] Optical rotation:α 20D -5.90°(c=0.235H2O) Example 14 Preparation of 5-methyl-(6S)-tetrahydrofolate monocholine salt Under argon, 2.61 g (4.611 mmol) of calcium levomefophosphate 3.8-hydrate was suspended in 60 ml of 95°C water. Next, an aqueous solution of filtered choline fluoride (obtained from choline chloride and silver fluoride) 568 mg (4.611 mmol) was added. The mixture was stirred at 95°C for 10 minutes, then cooled to room temperature. The precipitated calcium fluoride was filtered off using a high-flow pad, and the resulting clear solution (pH 6.5) was evaporated to dryness to obtain 2.568 g of the title compound. The crude product was immersed in 50°C ethanol, then cooled in an ice bath, filtered, and dried under vacuum at 60°C to obtain 1.67 g of the title compound. Analytical data: [Table 32] Optical rotation:α 20D +0.1° (c=1H2O) Example 15 Preparation of 5-formyl-(6S)-tetrahydrofolate diacetylcholine salt Under argon, 330.6 mg (0.5496 mmol) of calcium levofolinat pentahydrat was dissolved in 6 ml of 70°C water. Next, a filtered aqueous solution of 181.6 mg (1.099 mmol) of 30°C acetylcholine fluoride (obtained from acetylcholine chloride and silver fluoride) was added. After cooling to 0°C, the precipitated calcium fluoride was filtered off using a syringe filter, and the resulting clear solution (pH 6) was evaporated to dryness to obtain 407 mg of the title compound. Analytical data: [Table 33] Optical rotation:α 20D -3.4° (c=1H2O) Example 16 Preparation of 5-methyl-(6S)-tetrahydrofolate diacetylcholine salt Under argon, 331.8 mg (0.5862 mmol) of calcium levomefophosphate 3.8 hydrate was suspended in 8 ml of 70°C water. Next, a filtered aqueous solution of 193.6 mg (1.172 mmol) of acetylcholine fluoride (obtained from acetylcholine chloride and silver fluoride) at 50°C was added. After cooling to 0°C, the precipitated calcium fluoride was filtered off using a syringe filter, and the resulting clear solution (pH 6.5) was evaporated to dryness to obtain 433 mg of the title compound. Analytical data: [Table 34] Optical rotation:α 20D 17.04°(c=1H2O) Example 17 Preparation of 5-formyl-(6S)-tetrahydrofolate dibetamethyl ester salt Under argon, 50.6 mg (0.0736 mmol) of silver levofolinate was suspended in 2 ml of water. Next, a solution of 38.1 mg (0.1473 mmol) of betaine-methyl ester iodide was added to 1 ml of water, and the mixture was heated to 90°C while stirring. After cooling, the suspension was filtered using a syringe filter, and the resulting clear solution was evaporated and dried under vacuum at 50°C to obtain 56 mg of the title compound. Analytical data: [Table 35] Example 18 Preparation of 5-methyl-(6S)-tetrahydrofolate dibetamethyl ester salt Under argon, 520.1 mg (0.919 mmol) of calcium levomefophosphate 3.8 hydrate was suspended in 6 ml of 70°C water. Next, 291.8 mg (1.930 mmol) of filtered 20°C betaine methyl ester fluoride (obtained from betaine methyl ester iodide and silver fluoride) aqueous solution was added. After cooling to 0°C, the precipitated calcium fluoride was filtered off using a syringe filter. The resulting clear solution (pH 6.5) was evaporated and dried under vacuum at 40°C to obtain 682 mg of the title compound. Analytical data: [Table 36] Optical rotation:α 20D +10.5° (c=1H2O) Example 19 Preparation of 5-formyl-(6S)-tetrahydrofolate di2-dimethylaminoethanol Under argon, 58.0 mg (0.1225 mmol) of levofolinic acid was suspended in 0.5 ml of water and treated with 24.7 μl (0.2450 mmol) of 2-dimethylaminoethanol (Deanol). The mixture was stirred at room temperature (rt) until a clear solution was formed, evaporated, and dried under vacuum at 45°C to obtain 77 mg of the title compound. Analytical data: [Table 37] Optical rotation:α 20D -8.5° (c=1H2O) Example 20 Preparation of 5-formyl-(6S)-tetrahydrofolate mono-2-dimethylaminoethanol Under argon, 71.6 mg (0.1512 mmol) of levofolinic acid was suspended in 2 ml of water and 5 ml of methanol, and treated with 15.7 μl (0.1558 mmol) of 2-dimethylaminoethanol (Deanol). After sonication, the solution was immediately heated to 60°C to form a clear solution (pH 5). This solution was evaporated and dried under vacuum at 45°C to obtain 89.6 mg of the title compound. Analytical data: [Table 38] Optical rotation:α 20D -13.8° (c=1H2O) Example 21 Preparation of 5-methyl-(6S)-tetrahydrofolate mono-L-arginine salt Under argon, 82.0 mg (0.1785 mmol) of levomefolic acid was suspended in 10 ml of water and 20 ml of methanol. 31.1 mg (0.1785 mmol) of L-arginine was added, and the mixture was treated until a solution formed. This was evaporated, filtered by warm immersion with acetonitrile, and dried under vacuum at 50°C to obtain 109.7 mg of the title compound. Analytical data: [Table 39] Example 22 Preparation of 5-formyl-(6S)-tetrahydrofolate monoacetylcholine salt Under argon, 656.0 mg (1.090 mmol) of levofolinatocalcium pentahydratus was dissolved in 10 ml of 70°C water. Next, a filtered aqueous solution of 180.1 mg (1.090 mmol) of 30°C acetylcholine fluoride (obtained from acetylcholine chloride and silver fluoride) was added. After cooling to 0°C, the precipitated calcium fluoride was filtered off using a syringe filter, and the resulting clear solution (pH 6) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile and dried under vacuum at 50°C to obtain 640 mg of the title compound. Analytical data: [Table 40] Optical rotation:α 20D -7.7° (c=1H2O) Example 23 Preparation of 5-methyl-(6S)-tetrahydrofolate monoacetylcholine salt Under argon, 625.6 mg (1.105 mmol) of calcium levomefophosphate 3.8 hydrate was suspended in 12 ml of 70°C water. Next, 182.6 mg (1.105 mmol) of filtered 30°C aqueous solution of acetylcholine fluoride (obtained from acetylcholine chloride and silver fluoride) was added. The mixture was immediately heated to 60°C and then cooled again to 20°C. The precipitated calcium fluoride was filtered using a syringe filter, and the resulting clear solution (pH 6) was evaporated to dryness. The crude product was then filtered by warm immersion in acetonitrile and dried under vacuum at 50°C to obtain 637 mg of the title compound. Analytical data: [Table 41] Optical rotation:α 20D +29.7° (c=1H2O) The solubility of the selected folate was generally measured as follows: A quantity of salt corresponding to the expected solubility (between 1 and 100 mg) was provided, and each solvent was mixed while increasing the amount until a solution was formed. Highly soluble salts, in particular, may require longer processing times, as they can result in extremely viscous solutions. The solubility of two salts, 5-formyl-(6S)-tetrahydrofolate di-arginine salt and 5-methyl-(6S)-tetrahydrofolate di-arginine salt, in glycerol was tested in more detail. First, the di-arginine salts were finely ground in an agate mortar and pestle, then mixed with glycerin (50 mg (5% m / v) in 1 ml of solvent, under argon), and stirred with a magnetic stirrer. After stirring overnight, a clear solution was obtained. After confirming a solubility of 5% (m / v) prior to this, the same procedure was performed at twice the concentration to obtain a solution. Even at a concentration of 20% (m / v), the obtained solution was very viscous. For comparison, the solubility of the calcium salt of 5-methyl-(6S)-tetrahydrofolate (Gelpell) in glycerin was measured. The salt was clearly insoluble at 5% (m / v), 2.5% (m / v), and 1.6% (m / v). At 1.25% (m / v), a slightly milky solution was initially obtained, but it dissolved completely after stirring over the weekend. This confirmed previous data indicating that this calcium salt has a solubility of approximately 1%. The solubility of arginine (ground) in glycerin was observed to be good at 100 mg per 500 μl, and at 200 mg per 500 μl, a clear solution was obtained after stirring overnight and 36 hours. (Quality glycerin: 99.5%, assay 99.99%, water not specified. (Density: 1.26 g / ml)) All solubility data is expressed in mass % (m / m). Solubility was measured at 20°C. [Table 42] Quantitative HPLC measurement (20.00 mg of substance in 10.00 ml of H₂O₃) 2 Dissolve in O, inject 1.0 μl, at 280 nm. Table 43
Claims
1. An amorphous folate salt comprising a tetrahydrofolate anion and an organic cation, characterized in that the anion is 5-methyl-(6S)-tetrahydrofolate and the cation is an organic compound selected from the group consisting of diarginine, dicholine, and diacetylcholine.
2. The tetrahydrofolate anion is characterized by being 5-methyl-(6S)-tetrahydrofolate, and the organic cation is diarginine. ２ O Middle １ The H-NMR shift is, Table 1 The amorphous folate according to claim 1.
3. A pharmaceutical composition comprising at least one folate according to claim 1 or 2 and at least one pharmaceutically acceptable excipient.
4. A drug, food additive, or nutritional supplement for preventing and / or treating any deficiency or disorder that is positively affected by the administration of tetrahydrofolate, comprising at least one folate according to claim 1 or 2.
5. A method for preparing an amorphous folate according to claim 1 or 2, comprising the step of adding oxalic acid or, alternatively, a fluoride salt to an aqueous composition of an alkaline earth metal salt of folate.