Crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid
11377448 · 2022-07-05
Assignee
Inventors
- Rudolf Moser (Schaffhausen, CH)
- Viola Groehn (Dachsen, CH)
- Fritz Blatter (Reinach, CH)
- Martin Szelagiewicz (Basel, CH)
- Ruth Boehni Stamm (Stein Am Rhein, CH)
- Markus Ruettimann (Winterthur, CH)
Cpc classification
A61P25/28
HUMAN NECESSITIES
C07D475/04
CHEMISTRY; METALLURGY
International classification
Abstract
The present invention is directed to a crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid wherein the molar ratio of 5-methyl-(6S)-tetrahydrofolic acid to sodium is from 1:1.51 to 1:2.5 and/or hydrates and/or solvates thereof, as well as, a process of obtaining the same.
Claims
1. A crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid wherein the molar ratio of 5-methyl-(6S)-tetrahydrofolic acid to sodium is from 1:1.51 to 1:2.5 and/or a hydrate and/or a solvate thereof.
2. The crystalline salt of claim 1, wherein the molar ratio of 5-methyl-(6S)-tetrahydrofolic acid to sodium is from 1:1.75 to 1:2.25 and/or hydrates and/or solvates thereof.
3. The crystalline salt of claim 1, wherein the molar ratio of 5-methyl-(6S)-tetrahydrofolic acid to sodium is approximately 1:2 (in mol/mol).
4. The crystalline salt of claim 1, characterized in that the salt is the sodium salt of 5-methyl-(6S)-tetrahydrofolic acid and has a PXRD pattern with at least one characteristic peak (expressed in 2θ±0.3° 2θ(CuKα radiation)) at 3.2, 6.4, 7.8, 9.6, 12.7, 13.3, 13.9, 14.2, 14.7, 15.6, 16.3, 16.7, 17.2, 17.8, 18.2, 18.5, 19.3, 19.6, and 20.3.
5. The crystalline salt of claim 1, characterized in that the salt is the sodium salt of 5-methyl-(6S)-tetrahydrofolic acid and has a PXRD pattern with at least one characteristic peak (expressed in 2θ±0.3° 2θ (CuKα radiation)) at 3.2, 6.4, 7.8, 9.6, 12.7, 13.3, 13.9, 14.2, 14.7, 15.6, 16.3, 16.7, 17.2, 17.8, 18.2, 18.5, 19.3, 19.6, 20.0, 20.3, 20.7, 21.5, 22.0, 22.9, 23.5, 24.0, 24.6, 25.0, 25.4, 27.2 and 28.7.
6. The crystalline salt of claim 1, characterized in that the sodium salt of 5-methyl-(6S)-tetrahydrofolic acid has a PXRD pattern substantially as shown in
7. The crystalline according to claim 1, having at least 99% or more chemical and stereoisomerical purity.
8. A process for obtaining the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid according to claim 1 comprising the steps of: a) providing of 5-methyl-(6S)-tetrahydrofolic acid, optionally in a suitable solvent or a mixture of solvents; b) adding sodium hydroxide to the composition of step a); c) optionally adding a solvent, mixture of solvents and/or a co-salt former to the composition of step b), or adding the composition of step b) to a solvent, a mixture of solvents and/or a co-salt former; d) crystallizing; e) optionally adding more solvent or mixture of solvents; and f) isolating the obtained solid.
9. The process of claim 8, characterized in that the molar ratio of 5-methyl-(6S)-tetrahydrofolic acid and sodium hydroxide in step b) is in the range of from 1:1.51 to 1:5.
10. The process of claim 8, characterized in that the molar ratio of 5-methyl-(6S)-tetrahydrofolic acid and sodium hydroxide in step b) is in the range of from 1:1.9 to 1:3.
11. The process of claim 8, characterized in that solvent and/or mixtures mixture of solvents according to step a), c) and/or e) is selected from the group consisting of water, water-soluble alcohols, acetonitrile, tetrahydrofuran, acetone, methyl ethyl ketone, methyl isobutyl ketone, benzyl alcohol, or a mixture thereof.
12. The process of claim 8, characterized in that the co-salt former of step c) is an organic base with a pKa value from 6 to 11.
13. The process of claim 8, characterized in that the co-salt former of step c) is an organic base with a pKa value from 7 to 10.
14. The process of claim 8, characterized in that in step d) the temperature is at least 15° C.
15. The process of claim 8, characterized in that in step a), b), c) and/or d) seed crystals are added.
16. A pharmaceutical composition, food additive and/or vitamin comprising the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid according to claim 1 and optionally one or more acceptable excipients.
17. The pharmaceutical composition according to claim 16 in the form of tablets, capsules, oral liquid preparations, powders, lyophilisates, granules, lozenges, reconstitutable powders, injectable or infusable solutions or suspensions or suppositories.
18. The pharmaceutical composition according to claim 16 further comprising at least one additional therapeutic agent.
19. The pharmaceutical composition according to claim 16, which is a pharmaceutical composition for oral, parenteral, intramuscular, intraspinal, intrathecal, periodontal, topical or rectal administration.
20. A method which comprises adding the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid according to claim 1 to a drug or food additive.
21. A method comprising using the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid according to claim 1 in homocysteine lowering or in the treatment of anemia or neural tube defects, or in dietary management of low plasma folate, low red blood cell folate, low cerebrospinal fluid folate or low peripheral or central nervous system folate.
22. The process of claim 11, characterized in that a solvent and/or mixture of solvents according to step a), c) and/or e) is a water-soluble alcohol, selected from the group consisting of methanol, ethanol, isopropanol and n-propanol.
Description
BRIEF DESCRIPTION OF THE DRAWINGS
(1)
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EXPERIMENTAL
(9) Powder X-Ray Diffraction
(10) Stoe Stadi P equipped with a Mythen1K Detector; Cu-Kα1 radiation; standard measurement conditions: transmission; 40 kV and 40 mA tube power; curved Ge monochromator; 0.02°2θ step size, 48 s step time, 1.5-50.5°2θ scanning range; detector mode: step scan; 1°2θ detector step; standard sample preparation: 10 to 20 mg sample was placed between two acetate foils; sample holder: Stoe transmission sample holder; the sample was rotated during the measurement. All sample preparation and measurement was done in an ambient air atmosphere.
(11) Raman Spectroscopy
(12) FT-Raman spectra were recorded on a Bruker MultiRAM FT-Raman or a Bruker RFS 100 FT-Raman system with a near infrared Nd:YAG laser operating at 1064 nm and a liquid nitrogen-cooled germanium detector. 64 scans with a resolution of 2 cm.sup.−1 were accumulated in the range from 3500 to −50 cm.sup.−1; however, only data above 100 cm.sup.−1 are evaluated due to filter cutoff effects. Nominal laser powers are typically 100 or 300 mW.
(13) TG-FTIR
(14) Thermogravimetric measurements were carried out with a Netzsch Thermo-Microbalance TG 209 coupled to a Bruker FTIR Spectrometer Vector 22 (sample pans with a pinhole, N.sub.2 atmosphere, heating rate 10 K/min).
Example 1: Crystalline Sodium Salt of 5-methyl-(6S)-tetrahydrofolic acid Produced from Amorphous Disodium Salt of 5-methyl-(6S)-tetrahydrofolic acid (Seeding with Calcium Salt of 5-methyl-(6S)-tetrahydrofolic acid)
(15) 250.9 mg of amorphous 5-methyl-(6S)-tetrahydrofolic acid disodium salt were weighed into a 22 mL Supelco glass vial equipped with a magnetic stirrer bar. 2.0 mL of ethanol were added. The sample was sonicated for 1 minute and stirred at room temperature for 12 minutes. The gas phase of the vial was purged for 1 minute with nitrogen (about 100 mL/min). The sample was then stirred at 80° C. for 10 minutes. 3.0 mL of ethanol was added (in total 5.0 ml ethanol) and 0.5 mL of water were added (water activity about 0.50). The sample was sonicated for about 10 seconds at room temperature and stirred again at 80° C. for 5 minutes. The sample was seeded with about 1 mg of crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid and stirred at 80° C. for 3 minutes. An off-white suspension was formed and solid material that adhered to the glass wall of the vial was scraped off a spatula and a yellow suspension formed. The suspension was stirred at 80° C. for about 17 minutes and the solid material was again scraped from the glass wall using a spatula. The suspension was easy to stir. To the orange suspension, 3.0 mL of acetonitrile were added and the suspension was then stirred at 80° C. for 30 minutes. Hot filtration was carried out. The suspension was filtered using a fritted glass filter (porosity P4, Ø 1 cm) and the filter cake was air dried for 3 minutes. The filter cake was then transferred into weighing paper and further air dried for about 12 minutes at a relative humidity of about 40%. The solid product was 178 mg of a fine yellowish powder. Characterization by light microscopy and PXRD confirmed the crystalline nature of the solid. H-NMR spectroscopy confirmed the chemical integrity of 5-methyl-(6S)-tetrahydrofolic acid. The solid product shows a PXRD pattern as shown in
(16) TABLE-US-00001 TABLE 1 Powder X-ray diffraction data for crystalline sodium salt of Example 1 with 2-theta angles, d-spacing values in Ångstrom and qualitative intensity values as follows: vs = very strong, s = strong, m = medium, w = weak and vw = very weak. intensity ° 2-theta d-spacings [Å] (qualitative) 3.2 27.6 vs 6.4 13.8 vs 7.8 11.3 m 9.6 9.2 m 12.7 7.0 s 13.3 6.7 m 13.9 6.4 m 14.2 6.2 m 14.7 6.0 m 15.6 5.67 m 16.3 5.42 m 16.7 5.32 m 17.2 5.16 m 17.8 4.98 m 18.2 4.88 m 18.5 4.78 s 19.3 4.60 s 19.6 4.52 s 20.0 4.44 m 20.3 4.37 s 20.7 4.28 m 21.5 4.12 m 22.0 4.03 m 22.9 3.88 m 23.5 3.78 m 24.0 3.70 m 24.6 3.62 m 25.0 3.56 m 25.4 3.50 m
(17) TABLE-US-00002 TABLE 2 Raman data for the crystalline sodium salt of Example 1 with wavenumber in cm.sup.−1 and intensity values. It should be noted that the intensities vary with Laser power, sample amount and other factors. intensity wavenumber (cm.sup.−1) (arbitrary units) 3055 0.15 2929 0.32 1611 1.78 1582 0.49 1536 0.44 1483 0.30 1462 0.32 1418 0.22 1381 0.19 1329 0.44 1311 0.48 1276 0.40 1192 0.24 1021 0.15 949 0.14 875 0.28 835 0.14 776 0.15 651 0.55 621 0.24 479 0.17 363 0.15
Example 2: Crystalline Sodium Salt of 5-methyl-(6S)-tetrahydrofolic acid Produced from Amorphous Disodium Salt of 5-methyl-(6S)-tetrahydrofolic acid (Seeding with Crystalline Monosodium Salt of 5-methyl-(6S)-tetrahydrofolic acid)
(18) 1.2055 g of amorphous 5-methyl-(6S)-tetrahydrofolic acid disodium salt were weighed into a 40 mL Supelco glass vial equipped with a magnetic stirrer bar. 10.0 mL of ethanol/water 10:1 v/v were added. The suspension was stirred at room temperature for 8 minutes and seeded with 12 mg of crystalline monosodium salt of 5-methyl-(6S)-tetrahydrofolic acid. The suspension was stirred at room temperature for 5 minutes and the gas phase of the vial was purged for 1 minute with nitrogen (about 100 mL/min). An additional 10 mL of ethanol/water 10:1 v/v (water activity about 0.50) were added. The suspension was now easy to stir. It was sonicated for 1 minute and stirred at room temperature for 50 minutes. Microscopy of the suspension showed the presence of amorphous material. The gas phase of the vial was again purged for 1 minute with nitrogen (about 100 mL/min). The sample was then stirred at about 70° C. for 10 minutes. The sample was further stirred at about 70° C. for 10 minutes, seeded again with crystalline monosodium salt of 5-methyl-(6S)-tetrahydrofolic acid and stirred at about 70° C. for 8 minutes. A very weak suspension was formed. Stirring was continued at 70° C. for about 3 hours until a suspension formed that did not contain sticky amorphous solid as part of the mixture. As soon as an in-process control by light microscopy showed that the sample was predominantly crystalline hot filtration was carried out. The filter cake was air dried for 15 minutes. The fine, yellowish powder (0.8424 g) was examined by TG-FTIR. Since the mass loss was 15.4% (water and some ethanol) the sample was transferred into a 15 mL Supelco vial and further dried at room temperature and about 10 mbar for 20 minutes. About 800 mg of a fine, yellowish powder was obtained. Characterization by light microscopy and PXRD confirmed the crystalline nature of the solid. H-NMR spectroscopy confirmed the chemical integrity of 5-methyl-(6S)-tetrahydrofolic acid.
Example 3: Kinetic Solubility of Crystalline Sodium Salt of 5-methyl-(6S)-tetrahydrofolic acid
(19) 50 mg of crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid (according to Example 1) is weighed into a 4 ml glass vial with screw cap. Then 0.5 ml of purified/de-ionized water is added. The mixture is vigorously agitated at room temperature and briefly sonicated and a clear slightly yellow solution is readily obtained (within a few seconds). Thus the solubility is greater than 100 mg per 1 ml of water. The solution remains clear for several hours at r.t.
Reference Example 4: Kinetic Solubility of the Calcium Salt of 5-methyl-(6S)-tetrahydrofolic acid
(20) 27.9 mg of crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid (containing about 11% of water, thus corresponding to a dry weight of about 25 mg) are weighed into a 4 ml glass vial with screw cap. 2.535 ml of purified/de-ionized water is added to the solid using an adjustable volumetric pipette. The mixture is vigorously agitated at room temperature and briefly sonicated. No clear solution can be obtained and a fairly concentrated suspension persists. Thus the kinetic solubility measured as described here is smaller than 10 mg per 1 ml of water.
Example 5: Preparation of the Crystalline Disodium Salt of 5-methyl-(6S)-tetrahydrofolic acid Starting from 5-methyl-(6S)-tetrahydrofolic acid
(21) A mixture of 10 g 5-methyl-(6S)-tetrahydrofolic acid (assay 5-methyltetrahydrofolic acid 95.59% w/w), 20 g water and 1.74 g sodium hydroxide was heated in a glass vessel under a nitrogen atmosphere to 72° C. while stirring. At 72° C. 200 mL ethanol/5% v/v 2-propanol were added within 3.5 hours. Then 0.05 g of crystalline disodium salt of 5-methyl-(6S)-tetrahydrofolic acid were added as seeding material. At 72° C. additional 100 mL ethanol/5% v/v 2-propanol were added within 3 hours. The mixture was cooled to 25° C. within 14 hours and heated again to 72° C. The mixture was treated with ultrasonic to remove material which was sticking to the wall of the glass vessel, while the temperature decreased to 63° C. The solids were separated by filtration with nitrogen pressure in a closed filtration tube and washed with 100 mL ethanol/5% v/v 2-propanol. The solids were dried for 14 hours in vacuum at 36° C. to give 11.03 g of an off-white powder corresponding to 94.5% yield (assay 5-methyltetrahydrofolic acid 81.86% w/w). The solid product shows a PXRD pattern as shown in
Reference Example 6: Preparation of the Amorphous Monosodium Salt of 5-methyl-(6S)-tetrahydrofolic acid
(22) 6.0 g of crystalline 5-methyl-(6S)-tetrahydrofolic acid monosodium salt prepared according to EP 17164349.7 were added at room temperature to 600 g water. Solids were removed by suction filtration. The filtrate was frozen in a bath with liquid nitrogen and lyophilized in vacuum at 0.07 mbar. After 2 days the vacuum was replaced by nitrogen gas. 6.0 g amorphous 5-methyl-(6S)-tetrahydrofolic acid monosodium salt were obtained with a purity of 95.2% area, water content 8.8% w/w, assay sodium 4.5% w/w.
Example 7: Stability of the Crystalline Sodium Salt of 5-methyl-(6S)-tetrahydrofolic acid
(23) In order to compare the long-term stabilities of the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid, the compounds of the invention, to the long-term stability of the amorphous monosodium salt of 5-methyl-(6S)-tetrahydrofolic acid prepared according to Reference Example 6 and the crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid as prepared according to EP 1 044 975 B1, respective stability data has been generated at various temperatures and humidities.
(24) (a) Stability of the Crystalline Sodium Salt of 5-methyl-(6S)-tetrahydrofolic acid at 25° C./60% rh
(25) Amorphous monosodium salt of 5-methyl-(6S)-tetrahydrofolic acid prepared according to Reference Example 6, crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid, prepared according to literature procedures (EP 1 044 975 B1) and crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid, prepared as disclosed in Example 1, were stored at 25° C./60% rh. The content of 5-methyl-(6S)-tetrahydrofolic acid remaining in the samples was measured by HPLC at periodic intervals (% w/w). The results are shown in Table 1a and
(26) TABLE-US-00003 TABLE 1a Long-term stability of the crystalline sodium salt of 5- methyl-(6S)-tetrahydrofolic acid at 25° C./60% rh (% w/w) 5-methyl-(6S)-tetrahydrofolic acid (% w/w) 0 3 6 9 12 months months months months months amorphous 84.9 60.6 63.2 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline sodium 82.3 80.0 78.6 74.7 salt of 5-methyl- (6S)-tetrahydrofolic acid crystalline calcium 80.7 79.0 77.7 78.5 77.9 salt of 5-methyl- (6S)-tetrahydrofolic acid
(27) TABLE-US-00004 TABLE 1b Long-term stability of the crystalline monosodium salt of 5- methyl-(6S)-tetrahydrofolic acid at 25° C./60% rh (% rel.) 5-methyl-(6S)-tetrahydrofolic acid (% rel.) 0 3 6 9 12 months months months months months amorphous 100.0 71.3 74.5 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline sodium 100.0 97.3 95.5 90.8 salt of 5-methyl- (6S)-tetrahydrofolic acid crystalline calcium 100.0 97.9 96.3 97.3 96.5 salt of 5-methyl- (6S)-tetrahydrofolic acid
(28) TABLE-US-00005 TABLE 2 Long-term stability of the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid at 25° C./60% rh (major degradation product [MeFox]) Pyrazino-s-triazine derivative of 4α-hydroxy-5- methyl-THF (MeFox) (% w/w) 0 3 6 9 12 months months months months months amorphous 0.43 2.60 2.75 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline 0.07 1.01 1.31 1.42 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline calcium 0.20 0.84 0.90 0.85 0.86 salt of 5-methyl- (6S)-tetrahydrofolic acid
(b) Stability of the Crystalline Sodium Salt of 5-methyl-(6S)-tetrahydrofolic acid at 40° C./75% rh
(29) Amorphous monosodium salt of 5-methyl-(6S)-tetrahydrofolic acid prepared according to Reference Example 6, crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid, prepared according to literature procedures (EP 1 044 975 B1) and crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid, prepared as disclosed in Example 1, were stored at 40° C./75% rh. The content of 5-methyl-(6S)-tetrahydrofolic acid remaining in the sample was measured by HPLC at periodic intervals (% w/w). The results are shown in Table 3a and
(30) TABLE-US-00006 TABLE 3a Long-term stability of the crystalline sodium salt of 5- methyl-(6S)-tetrahydrofolic acid at 40° C./75% rh (% w/w) 5-methyl-(6S)-tetrahydrofolic acid (% w/w) 0 3 6 9 12 months months months months months amorphous 84.9 61.6 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline sodium 82.3 78.6 77.3 77.9 salt of 5-methyl- (6S)-tetrahydrofolic acid crystalline calcium 80.7 78.5 78.0 78.8 77.8 salt of 5-methyl- (6S)-tetrahydrofolic acid
(31) TABLE-US-00007 TABLE 3b Long-term stability of the crystalline monosodium salt of 5- methyl-(6S)-tetrahydrofolic acid at 40° C./75% rh (% rel.) 5-methyl-(6S)-tetrahydrofolic acid (% rel.) 0 3 6 9 12 months months months months months amorphous 100.0 72.6 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline sodium 100.0 95.6 94.0 94.7 salt of 5-methyl- (6S)-tetrahydrofolic acid crystalline calcium 100.0 97.2 96.6 97.6 96.3 salt of 5-methyl- (6S)-tetrahydrofolic acid
(32) TABLE-US-00008 TABLE 4 Long-term stability of the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid at 40° C./75% rh (major degradation product [MeFox]) Pyrazino-s-triazine derivative of 4α-hydroxy-5- methyl-THF (MeFox) (% w/w) 0 3 6 9 12 months months months months months amorphous 0.43 2.79 monosodium salt of 5-methyl-(6S)- tetrahydrofolic acid crystalline sodium 0.07 1.52 1.69 1.51 salt of 5-methyl- (6S)-tetrahydrofolic acid crystalline calcium 0.20 1.01 0.96 0.93 0.95 salt of 5-methyl- (6S)-tetrahydrofolic acid
(33) Tables 1 to 4 with the stability data of crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid as disclosed in in the present invention clearly shows that
(34) i) there is a remarkable difference in the stability of crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid compared to the amorphous monosodium salt crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid and
(35) ii) the crystalline sodium salt of 5-methyl-(6S)-tetrahydrofolic acid is showing a comparable stability over a long period of time to the crystalline calcium salt of 5-methyl-(6S)-tetrahydrofolic acid.