METHOD FOR PRODUCING PURE L-HERCYNINE

Abstract

The invention relates to a method for the purification of L-hercynine. Said method for the purification of L-hercynine, from a reaction mixture resulting from the reaction of L-histidine, in controlled pH conditions, with a methylating agent Me X in a polar solvent or a mixture of polar solvents, at room temperature, is characterised in that it comprises at least one step of separating the organic products from the inorganic salts formed during the reaction by electrodialysis. This method allows the L-hercynine losses to be limited during the purification.

Claims

1. A method for purifying L-hercynine, or its enantiomer, D-hercynine, or its racemic mixture, or one of its isotopically marked derivatives from a reaction mixture resulting from the reaction of L-histidine, or its enantiomer D-histidine or its racemic mixture, or one of its isotopically marked derivatives or one of its a-N-methylated derivatives, under controlled pH conditions, with a methylation agent MeX in a polar solvent or a mixture of polar solvents, at room temperature, characterized in that it comprises at least one step for separating the organic products from the inorganic salts formed during the reaction by electrodialysis; and in that the electrodialysis is conducted on a reaction medium adjusted to a pH comprised between 8.5 and 9.5, at the beginning of the electrodialysis.

2. The purification method according to claim 1, characterized in that it comprises an additional step for recrystallization of the L-hercynine, or its enantiomer, D-hercynine, or its racemic mixture, or one of its isotopically marked derivatives or one of its methylated derivatives, to eliminate the organic impurities.

3. A method for the industrial-scale production of L-hercynine, or its enantiomer, D-hercynine, or its racemic mixture, or one of its isotopically marked derivatives, or one of its a-N-methylated derivatives, characterized in that it comprises the following steps: reacting the L-histidine, or its enantiomer D-histidine or its racemic mixture, or one of its isotopically marked derivatives or one of its a-N-methylated derivatives, under controlled pH conditions, with a methylation agent MeX in a polar solvent or a mixture of polar solvents, at room temperature; followed by separating the organic products from the inorganic salts formed during the reaction by electrodialysis; and in that the electrodialysis is done on a reaction medium adjusted to a pH comprised between 8.5 and 9.5, upon starting the electrodialysis; followed by re-crystallizing the L-hercynine to eliminate the organic impurities.

4. The method according to claim 1, characterized in that the electrodialysis is done on a reaction medium adjusted to about 9, at the beginning of the electrodialysis.

5. The method according to claim 1, characterized in that the methylating agent MeX is chosen from among methyl halogenide, methyl sulfate, methyl carbonate or methyl or trifluoromethyl or tosyl sulfonate.

6. The method according to claim 1, characterized in that the polar solvent or the mixture of polar solvents is chosen from among water; a C1—C6 alcohol, for example methanol, ethanol, propanol or propanol-2, butanol; ethyl acetate. One particular polar mixture is a water/alcohol, water/propanol or propanol-2, methanol/ethyl acetate mixture.

7. The method according to claim 1 one of the claims 1 to 6, characterized in that the controlled pH conditions are chosen to result directly in a tri-methylation of the L-histidine or its D-histidine enantiomer or its racemic mixture, or one of its isotopically marked derivatives or one of its a-N-methylated derivatives, or indirectly from the di-methylated derivative.

8. The method according to claim 1, characterized in that the controlled pH conditions are chosen in the interval pH=9-13 by adding an inorganic base such as sodium hydroxide, potassium or lithium hydroxide.

9. The method according to claim 1, characterized in that the L-Hercynine-d9 is prepared from a reaction mixture resulting from the reaction of the L-Histidine with a methylation-d3 agent, in particular iodomethane-d3.

10. L-Hercynine-d9, with formula ##STR00012## in particular as obtained using the method according to claim 1.

Description

DESCRIPTION OF THE FIGURES

[0063] FIG. 1: Chromatogram (HPLC) of the L-hercynine obtained in example 2 on an industrial scale after desalination and recrystallization (ELSD Universal detection)

[0064] FIG. 2: Chromatogram (HPLC) of the L-hercynine-d9 obtained in example 3 after desalination and recrystallization (ELSD Universal detection)

EXAMPLES

[0065] The examples below, as well as the figures, are provided simply as an illustration and cannot in any way limit the scope of the invention, but are instead an integral part of the invention, in their general means.

[0066] In the examples described below, all percentages are given by weight, the temperature is room temperature or is given in degrees Celsius, and the pressure is the atmospheric pressure, unless otherwise stated. Room temperature refers to a temperature comprised between 20° C. and 35° C.

[0067] The reagents used are commercially available from international suppliers such as Sigma-Aldrich France (France), Alfa Aesar, Fisher Scientific, TCI Europe, Bachem (Switzerland), except the following compounds, which have been produced according to the cited protocol: N,N-Dimethyl-L-histidine hydrochloride hydrate (Reinhold, V. et al.; J. Med. Chem.; 1968; 11; 258-260).

[0068] The desalination was done by electrodialysis using cells made up of alternating anion-selective (for example, PC SA) and cation-selective (for example, PC MV) membranes in particular available from PC Cell (Germany), using desalination units like the ED 8002-001 unit (micro-unit for scale up to 10 g), the B-ED 1-3/ED 200 unit (up to 200 g) and the P20 unit (industrial scale), also available from PC Cell (Germany).

[0069] The NMR-.sup.1H analyses were recorded at 400 MHz in D.sub.2O or a D.sub.2O/DCI mixture, using the HOD signal (4.79 ppm) as internal reference. The chemical displacements are noted in ppm, and the multiplicity of the signals indicated by the following symbols: s (singlet), d (doublet), t (triplet), q (quartet), and m (multiplet). The coupling constraints are noted in Hertz (Hz). The NMR-.sup.13C analyses are recorded at 75 MHz in the D.sub.2O. The mass analyses are obtained by atmospheric pressure chemical ionization (APCI-MS). The melting points were measured with a device by the company Stuart Scientific. The HPLC analyses were done on an Acquity (Waters) device, using a C8 250x4.6 (5 μm) column. The mobile phase used is a water/acetonitrile mixture (98:2) in 12 min. and a flow rate of 0.6 mL/min. The detection is done with an ELSD (Sedere) universal detector.

I. EXAMPLES ACCORDING TO THE INVENTION OF THE METHOD FOR PRODUCING PURE COMPOUNDS

[0070] The following examples illustrate a new, simple and reproducible method for producing pure L-hercynine, or its enantiomer, D-hercynine, or its racemic mixture, or one of its isotopically marked derivatives that is based on a method for separating inorganic salts, present in the reaction medium, by electrodialysis of an aqueous solution adjusted to pH=9, irrespective of the methylation mode used for the L-histidine or one of its methylated derivatives, followed by purification via recrystallization.

Example 1 of the invention

Isolation of the Pure L-Hercynine from a Reaction Mixture Obtained by Tri-Methylation of the L-Histidine

[0071] ##STR00006##

[0072] 275 g (1.3 mol) of the hydrochloride hydrate of the L-histidine is solubilized in 2.7 L of a water/methanol mixture (2:1). The pH is adjusted to 9-13 with a 3M potassium hydroxide solution. Next, 645 g (4.55 mol) of iodomethane is added under agitation, and the mixture thus obtained is agitated for 18 hours at 25-35° C. while keeping the pH between 9-13. Next, the mixture is neutralized by adding 37% aqueous HC1.

[0073] After evaporation of the methanol, the pH of the aqueous solution, containing the raw product in mixture with the potassium chloride and iodide salts, is adjusted to 9 with a solution of NaOH (30%), then the solution is desalinated by electrodialysis until a conductivity of 50-100 μS is obtained. A test for the presence of halogen with silver nitrate, done on a withdrawn sample, confirms the absence of chloride and iodide ions.

[0074] The aqueous solution, obtained after electrodialysis, is dry evaporated. The raw product is purified by recrystallization in aqueous isopropanol, and the L-hercynine is obtained after filtration and drying (192 g; 73%) in the form of a white powder. [0075] [α]D=+44.5° (c=1.0; 5N HC1) [0076] pF: 238° C. (dec.) [0077] NMR-.sup.1H (D.sub.2O, 400 MHz):β (ppm)=3.24 (m+s, 11H), 3.90 (dd, J=10 Hz, J=5 Hz, 1H), 6.98 (s, 1H), 7.69 (s, 1H). [0078] NMR-.sup.1H (D.sub.2O/DCI, 400 MHZ):β (ppm)=3.31 (s, 9H), 3.44 (dd, J=14 Hz, J=12 Hz, 1H), 3.55 (dd, J=14 Hz, J=4 Hz, 1H), 4.13 (dd, J=12 Hz, J=4 Hz, 1H), 7.37 (s, 1H), 8.66 (s, 1H). [0079] NMR-.sup.13C (D.sub.2O, 75 MHz):β (ppm)=25.7; 52.4; 78.9; 116.7; 132.3; 136.5; 171.2. HPLC-MS (AP+): m/z=198 (MH+) [0080] Purity by HPLC (ELSD detection): 100%

Example 2 of the Invention

Isolation of the Pure L-Hercynine from a Reaction Medium Obtained by Tri-Methylation of the L-Histidine on the Industrial Scale

[0081] ##STR00007##

[0082] 48.0 kg (229 mol) of the hydrochloride hydrate of the L-histidine is solubilized in 475 liters of a water/methanol mixture (2:1). The pH is adjusted to 9-13 with a 10% sodium hydroxide solution. Next, 113.8 kg (801 mol) of iodimethane is added under agitation, and the mixture thus obtained is agitated for 18 hours at 25-35° C. while keeping the pH between 9-13. Next, the mixture is neutralized by adding 37% aqueous HC1.

[0083] After evaporation of the methanol, the pH of the aqueous solution, containing the raw product in mixture with the sodium chloride and iodide salts, is adjusted to 9 with a solution of NaOH (30%), then this solution is desalinated by electrodialysis until a conductivity of 50-100 μS is obtained. A test for the presence of halogen with silver nitrate, done on a withdrawn sample, confirms the absence of chloride and iodide ions.

[0084] The desalinated aqueous solution, obtained after electrodialysis, is concentrated. After recrystallization in aqueous isopropyl, the L-hercynine is obtained after filtration and drying (32.2 kg; 72%) in the form of a white powder. [0085] [α]D:+44° (c=1.0; 5N HC1) [0086] pF: 242° C. (dec.)

[0087] The NMR-.sup.1H and NMR-.sup.13C and mass spectrums are identical to those of the L-hercynine obtained in example 1.

[0088] Purity by HPLC (ELSD detection): 99.2% (see FIG. 1).

Example 3 of the Invention

Isolation of the Pure L-Hercynine-d9 From the Reaction Mixture Obtained by Tri-Methylation of the L-Histidine

[0089] ##STR00008##

[0090] 31.4 g (150 mmol) of the hydrochloride hydrate of the L-histidine is solubilized in 300 mL of a water/methanol mixture (2:1). The pH is adjusted to 9-13 with a 3M sodium hydroxide solution. Next, 86.9 g (600 mmol) of iodomethane-d3 is gradually added under agitation, while keeping the pH between 9-13. After 18 hours of agitation at 30° C., the mixture thus obtained is neutralized by adding 37% aqueous hydrochloric acid.

[0091] After evaporation of the methanol, the pH of the aqueous solution, containing the raw product in mixture with the sodium chloride and iodide salts, is adjusted to 9 with a NaOH solution (30%), then the solution is desalinated by electrodialysis until obtaining a conductivity of 50-100 μS. A test for the presence of halogen with silver nitrate, done on a withdrawn sample, confirms the absence of chloride and iodide ions.

[0092] The aqueous solution, obtained after electrodialysis, is dry evaporated. The obtained raw product is purified by recrystallization in the aqueous isopropanol, and the L-Hercynine-d9 is obtained after filtration and drying (23.8 g; 77%) in the form of a white powder. [0093] NMR-.sup.1H (D.sub.2O/DCI, 400 MHz):β (ppm)=3.14 (m, 2H); 3.83 (dd, J=10 Hz, J=5 Hz, 1H); 6.92 (s, 1H), 7.62 (s, 1H). [0094] HPLC-MS (AP+): m/z=207 (MH+). [0095] Purity by HPLC (ELSD detection): 100% (see FIG. 2).

Example 4 of the Invention

Isolation of the Pure L-Hercynine from a Reaction Mixture Obtained by Quaternization of the N,N-Dimethyl-Histidine with Iodomethane

[0096] ##STR00009##

[0097] The N,N-dimethyl-histidine is quaternized as described by Reinhold, V. et al.; J. Med. Chem.; 1968; 11; 258-260. In short, 48.5 g (200 mmol) of the hydrochloride hydrate of the N,N-dimethyl-histidine is solubilized in 1 liter of methanol. The pH is adjusted to 9 with a 20% ammonia aqueous solution. Next, 37.3 g (16 mL, 1.3 equivalents) of iodomethane is added under agitation. The solution thus obtained is agitated for 18 hours at 30° C. (room temperature). After evaporation of the methanol, the raw product, in the reaction mixture containing the ammonium chloride and iodide salts (88 g), is solubilized in 500 mL of water, the pH is adjusted to 9 with a NaOH (30%) solution, then the solution is desalinated by electrodialysis until a conductivity of 43 microsiemens (μS) is obtained. A test with silver nitrate, making it possible to estimate the presence of halogenide ions, done on a sample taken from the aqueous solution, confirms the absence of chloride and iodide ions.

[0098] The aqueous solution, obtained after electrodialysis, is dry evaporated. The raw product obtained is purified by recrystallization, and the L-hercynine is obtained after filtration and drying (36.6 g; 93%) in the form of a white powder. [0099] [α]D=+44° (c=1.0; 5N HC1) [0100] pF: 241° C. (dec.) [0101] NMR-.sup.1H (D.sub.2O, 400 MHz):β (ppm)=3.16 (m+s, 11H), 3.85 (dd, J=10 Hz, J=5 Hz, 1H), 6.93 (s, 1H), 7.64 (s, 1H). [0102] NMR-.sup.13C (D.sub.2O, 75 MHz):β (ppm)=25.7; 52.4; 78.9; 116.7; 132.3; 136.5; 171.2. [0103] HPLC-MS (AP+): m/z=198 (MH+)

Example 5 of the Invention

Isolation of the Pure D-Hercynine from a Reaction Mixture Obtained by Quaternization of the D-N,N-Dimethyl-Histidine with Iodomethane

[0104] ##STR00010##

[0105] The D-N,N-dimethyl-histidine is quaternized by analogy with example 4 describing the synthesis and purification of the L-hercynine. In short, 21.3 g (90 mmol) of the hydrochloric hydrate of the D-N,N-dimethyl-histidine is solubilized in 0.45 liters of methanol. The pH is adjusted to 9 with a 20% ammonia aqueous solution. Next, 15.3 g (6.7 mL, 1.2 equivalents) of iodomethane is added under agitation. The solution thus obtained is agitated for 18 hours at 25-30° C. After evaporation of the methanol, the raw product, in the reaction mixture containing the ammonium chloride and iodide salts, is solubilized in 300 mL of water. After adjustment of the pH of the solution to 9 with a NaOH (30%) solution, the mixture is desalinated by electrodialysis until a conductivity of 70 μS is obtained. A silver nitrate test, making it possible to estimate the presence of halogenide ions, done on a sample taken from the aqueous solution, confirms the absence of chloride and iodide ions.

[0106] The aqueous solution, obtained after electrodialysis, is dry evaporated. The raw product obtained is purified by recrystallization with a methanol/ethyl acetate mixture, and the L-hercynine is obtained after filtration and drying (16.7 g; 93%) in the form of a white powder. [0107] [α]D: -44° (c=1.0; 5N HC1) [0108] The NMR-.sup.1H and NMR-.sup.13C and mass spectrums are identical to those of the L-hercynine obtained in example 1.

Example 6 of the Invention

Isolation of the Pure L-Hercynine from a Reaction Mixture Obtained by Quaternization of the N,N-Dimethyl-Histidine with the Methyl Sulfate

[0109] ##STR00011##

[0110] The N,N-dimethyl-histidine is quaternized by analogy with the procedure described by Reinhold, V. et al.; J. Med. Chem.; 1968; 11; 258-260, replacing the iodomethane with methyl sulfate. In short, 4.85 g (20 mmol) of hydrochloride hydrate of the N,N-dimethyl-histidine is solubilized in 100 mL of methanol. The pH is adjusted to 9 with a 20% ammonia aqueous solution. Next, 3.03 g (2.28 mL, 24 mmol, 1.2 equivalents) of methyl sulfate is added under agitation. The solution thus obtained is agitated for 18 hours at 30° C. The NMR-.sup.1H analysis of a sample indicates that the conversion is not complete. 1.0 g (0.75 mL, 8 mmol, 0.4 equivalents) of methyl sulfate is added to the reaction mixture. After agitation for 3 hours, the solvent is evaporated, and the raw product, in the reaction mixture containing the ammonium chloride and iodide salts, is solubilized in 50 mL of water. After adjusting the pH to 9, desalination by electrodialysis is done until a conductivity of 50 μS is obtained, a silver nitrate test making it possible to estimate the presence of halogenide ions, done on a sample taken from the aqueous solution, and an analysis by HPLC-ELSD confirms the absence of salts.

[0111] The aqueous solution, obtained after electrodialysis, is dry evaporated. After recrystallization of the raw product with a methanol/ethyl acetate mixture, the L-hercynine is obtained after filtration and drying (3.0 g; 76%) in the form of a white powder.

[0112] The NMR-.sup.1H spectrum and the rotatory power are identical to those obtained for the isolated product in example 1.

II. COMPARATIVE EXAMPLES OF THE ELECTRODIALYSIS METHOD

Comparative Example 1

Evaluation of the Separation of the Inorganic Salts from the L-Hercynine at pH 7 and pH 8

[0113] A) 2000 mL of an aqueous solution containing 113 g (0.57 mol) L-hercynine, 300 g (3.5 equiv.) of sodium iodide and 33 g (1 equiv.) of sodium chloride (1.3 mol) at pH 7 and a conductivity of 70 mS/cm are placed in the “diluent” compartment of an electrodialysis device, for example the B-ED 1-3/ED 200 unit, equipped with an ED 200 cell. 2000 mL of demineralized water is placed in the “concentrate” compartment. 8000 mL of an aqueous solution of sodium sulfate (0.5-1%) is used as electrolyte. The circulation flow rate of all of the solutions is 100/120 L/h, by applying a voltage of 10-15 V, until the conductivity of the “diluent” is <0.1 mS/cm. A test for the presence of halogen with the silver nitrate, done on a withdrawn sample, confirms the absence of chloride and iodide ions. The “diluent” contains 86 g of L-hercynine, corresponding to a loss of 24%.

[0114] B) 2000 mL of an aqueous solution containing 113 g (0.57 mol) of L-hercynine, 300 g (3.5 equiv.) of sodium iodide and 33 g (1 equiv.) of sodium chloride (1.3 mol) at pH 8 and a conductivity of 75 mS/cm is electrodialyzed (as described in A) up to a conductivity of the “diluent” of 0.02 mS/cm. A test for the presence of halogen with silver nitrate, done on a withdrawn sample, confirms the absence of chloride and iodide ions. The diluent contains 98.3 g of L-hercynine, corresponding to a loss of 13%.

Comparative Example 2

Evaluation of the Separation of the Inorganic Salts from the L-Hercynine at pH 7 (Comparative) and pH 9 (According to the Invention)

[0115] A) The reaction mixture obtained by tri-methylation of the L-histidine, as described in example 2 of the invention, after neutralization with the hydrochloric acid HC1, containing 5% (w/w) of the hercynine mixed with reaction sub-products, as well as the sodium chloride and iodide salts, is desalinated in portion in 2000 mL by electrodialysis as described in comparative example 1A, until a conductivity of 50-100 μS is obtained. The L-hercynine content is assayed by HPLC. The procedure has been applied on 5 lots numbered 1 to 5, and the L-hercynine losses (from 22-25%) are listed in table 1.

[0116] B) Under the same conditions, with the exception that the pH is adjusted to 9, according to the invention, with 30% sodium hydroxide in the “diluent” compartment upon starting the electrodialysis, 5 lots numbered 6 to 10 are desalinated until obtaining a conductivity of 50-100 μS. The L-hercynine content in the “diluent” is assayed by HPLC. The procedure was applied on 5 lots, and the L-hercynine losses, going from only 1.5-2.5%, are listed in Table 1.

TABLE-US-00001 TABLE 1 L-hercynine losse Lot 1 Lot 2 Lot 3 Lot 4 Lot 5 Example loss 24% 22% 25% 22% 23% 2A: pH = 7 Lot 6 Lot 7 Lot 8 Lot 9 Lot 10 Example loss 2.5% 2.3% 1.9% 2% 1.5% 2B: pH = 9

[0117] Comparative examples 1 and 2 therefore indeed show that the losses during electrodialysis can be limited very significantly by adjusting the pH of the aqueous solution to 8.5-9.5, ideally to around 9, at the beginning of the electrodialysis, according to the invention.