PROCESS FOR THE PREPARATION OF N<G>-MONOMETHYL-L-ARGININE HYDROCHLORIDE

Abstract: The present invention relates to a process for the preparation of N<G>-monomethyl-L-arginine hydrochloride comprising reaction of a compound of formula (I) or an acid addition salt thereof, wherein L is a leaving group; with L-ornithine or an acid addition salt thereof.

Description: PROCESS FOR THE PREPARATION OF N c -MONOMETHYL-L-ARGININE HYDROCHLORIDE The present invention relates to a process for the preparation of N G - monomethyl-L-arginine hydrochloride (L-NMMA hydrochloride). A process for preparing N G -monomethyl-L-arginine and related compounds has been described in WO94/26701. On a large scale, this process suffers from the drawback that L-NMMA has to be isolated by column chromatography. Preparation of L-NMMA hydrochloride is also described in WO94/02453. The methods described in this publication would, however, be unsuitable for a commercial process. An improved synthesis of N G -allyl-(L)-arginine is described by Benatowicz et al in Synthetic Communications, 22 (5), 657-661 (1993). This process used fiavianic acid and ion exchange chromatography to isolate the product. We have now discovered a new process for the preparation of L-NMMA hydrochloride where the product can be isolated directly from the reaction mixture without the need for chromatography. The new process also results in higher yields and an improved impurity profile of the product. Accordingly, the present invention provides a process for the preparation of N G - monomethyl-L-arginine hydrochloride comprising: (i) reaction of a compound of formula (I) or an acid addition salt thereof, preferably a hydrochloride salt thereof, wherein L is a leaving group; with L-ornithine or an acid addition salt thereof, preferably a hydrochloride salt thereof, in a suitable polar solvent, for example, water, optionally in the presence of a base followed by: (ii) acidification to pH 4 to 5 with hydrochloric acid; then (iii) addition of an alcoholic solvent, such as ethanol or industrial methylated spirit; then (iv) isolation of N G -monomethyl-L-arginine hydrochloride. In the compound of formula (I), L is a leaving group as would be understood by a person skilled in the art. Most suitably, L is the group: wherein R 1 , R 2 , and R 3 are independently selected from hydrogen, halo, C^alkyl, C^alkoxy, and aryl, including fused aryl (ie where either R 1 and R 2 or R 2 and R 3 together with the carbon atoms to which they are attached form an aryl group fused to the pyrazole). However, L may also be C^alkoxy or C^cycloalkoxy. In the most preferred aspect of the invention, L is unsubstituted pyrazole. In the definition of L the term "aryl" means a 5 to 10 membered mono- or bi- cyclic aromatic system optionally including 1 to 3 heteroatoms selected from nitrogen, sulphur, and oxygen wherein the aromatic system is optionally substituted by 1 to 5 groups selected from halo, C^alkyl, and C^alkoxy. Suitable examples of such aryl substituents include phenyl and naphthyl. In the definition of L, the term "halo" means fluoro, chloro, bromo or iodo. The base used in step (i) may be any inorganic or organic base which liberates the free form of the compound of formula (I) and/or of L-omithine and which, in the presence of hydrochloric acid, forms a by-product which is soluble in the solvent mixture produced in step (iii) of the reaction. For economy and convenience, the most preferred base is lithium hydroxide. However, alkyl or aromatic amines such as imidazole may also be suitable. As would be understood by the skilled person, where the free forms of both the compound of formula (I) and of the L-ornithine are used, it may be possible to effect the reaction without adding a further base, otherwise the base is required. According to a preferred aspect, the present invention provides a process for the preparation of N G -monomethyl-L-arginine hydrochloride comprising: (i) reaction of pyrazole carboxamidine hydrochloride, N CH J NH NH .HCl with L-ornithine hydrochloride, in a suitable polar solvent, for example, water, in the presence of lithium hydroxide followed by: (ii) acidification to pH 4 to 5 with hydrochloric acid; then (iii) addition of an alcoholic solvent, such as ethanol or industrial methylated spirit; then (iv) isolation of N G -monomethyl-L-arginine hydrochloride. During Step (i), the reaction may proceed at high pH, such as pH 9 to 12; however it is preferable to maintain a pH in the range 10 to 11, for example between 10 and 10.5 by addition of acid, for example concentrated hydrochloric acid. The reaction is suitably carried out at a non-extreme temperature, such as 0 β C to 70°C, for example 0°C to 50°C, most preferably 10 to 30°C, most conveniently at ambient temperature. During step (ii), the hydrochloric acid is preferably added to give a pH of around 4.5 such that the mono-hydrochtoride salt is obtained in optimal yield. In step (iii), the purpose of the alcoholic solvent is to act as a solvent for the by¬ products of the reaction and as a non-solvent for the product N G -monomethyl-L- arginine hydrochloride. Therefore, the invention includes within its scope, any variation of step (iii), for example, substitution of the alcoholic solvent which fulfils this purpose. In step (iv) the product L-NMMA hydrochloride may be isolated directly from the reaction mixture by crystallisation, for example by adding seed crystals of L- NMMA hydrochloride and stirring the reaction mixture at a non-extreme temperature, such as -5°C to 30°C most conveniently at ambient temperature. The resultant product may optionally be further purified, for example by recrystallisation from a suitable solvent or mixture of solvents. As would be clear to a person skilled in the art, the compound of formula (I) used in the process of present invention may be replaced by methylcyanamide and this variation forms a further embodiment of the present invention. In a further embodiment, the present invention provides the novel intermediates of formula (I), such as pyrazole carboxamidine and salts thereof, in particular pyrazole carboxamidine hydrochloride. The intermediates of formula (I) are commercially avavilable or may be prepared from commercially available starting materials by standard methods of chemistry, for example, by reaction of the appropriate pyrazole with methyl cyanamide. Synthetic Example (a) Preparation of methylcyanamide Cyanogen bromide (21.2g) was dissolved in tetrahydrofuran (THF) (150ml) with sodium carbonate (42.4g) present. The whole was stirred and cooled to between -10 to 20°C and over this range of temperature, methylamine (2M solution in THF, 100ml) was added over 10 minutes. The stirred reaction mixture was then left to react for 2 hours at -15 to 20°C, then allowed to warm to 10°C before filtering off the solid sodium salts to give a clear solution of methylcyanamide in THF. (b) Preparation of 1 H-pyrazole-1-N-methylcarboxamidine hydrochloride Pyrazole (13.62g) was added to methylcyanamide solution (250ml) prepared as described in part (a) and then a hydrogen chloride solution (4M in dioxane 53ml) added. Upon addition of the hydrogen chloride a faint white precipitate appeared momentarily then dissolved. No significant exotherm was noted. The whole was heated with stirring at reflux for 2 hours forming a yellow liquid layer. The reaction mixture was then allowed to cool with stirring and a few crystals of presolidified product added. A cake of product resulted, this was broken up and stirred to give an off white solid/liquid mixture. To this was added diethyl ether (50ml) with stirring, then the whole was filtered to give an off white waxy solid. The solid was dried in vacuo at 50°C to give 22.6g of a cream solid product, mp 153-154°C. NMR analysis showed it to contain ca 5% pyrazole. 1 H NMR (DMSO-d 6 ), δppm : 3.55 (s, 3); 6.78 (s, 1); 8.09 (s, 1); 8.9 (s, 1). (c) Preparation of N G --monomethvl-L-arαinine hydrochloride 1H-Pyrazole-1-N-methylcarboxamidine hydrochloride (12.85g), prepared as described in part (b), and L-ornithine hydrochloride (8.43g) were mixed in water, pH 2 to 3. Lithium hydroxide monohydrate (5.45g) was added to give a pH of 10.1. A cream suspension formed which changed to a yellow solution after 1 to 2 hours and a pH of ca 11. The pH was brought down and maintained between 10 and 10.5 with concentrated hydrochloric acid. After 3-4 days stirring at room temperature between this pH range no ornithine was detected by t.l.c. silica (0.88NH 3 /MeOH 15:35). The reaction mixture was brought to pH 4.5 with concentrated hydrochloric acid (-5ml), then ethanol (200ml) added. A few seed crystals of N G -monomethyl-L- arginine hydrochloride were added and the whole stirred at room temperature for 3 days giving a white crystalline precipitate. This was filtered off to give 6g of the title product, mp 219-221°C with gas evolution. 1 HNMR (D 2 0), δ ppm : 1.6 (m, 2); 1.82 (q, 2); 2.74 (s, 3); 3.15 (t, 2); 3.7 (t, 1).

Claims: C AIMS A process for the preparation of N G -monomethyl-L-arginine hydrochloride comprising: (i) reaction of a compound of formula (I): or an acid addition salt thereof, wherein L is a leaving group; with L-ornithine or an acid addition salt thereof, in a suitable polar solvent, optionally in the presence of a base followed by: (ii) acidification to pH 4 to 5 with hydrochloric acid; then (iii) addition of an alcoholic solvent; then (iv) isolation of N G -monomethyl-L-arginine hydrochloride. 2. A process according to claim 1 wherein the compound of formula (I) is or an acid addition salt thereof, wherein R , R 2 , and R 3 are independently selected from hydrogen, halo, C^alkyl, C^alkoxy, and aryl, including fused aryl. A process accordingly to claim 1 or 2 wherein the compound of formula (I) is or an acid addition salt thereof. 4. A process according to any of claims 1 to 3 wherein the compound of formula (I) is in the form of a hydrochloride salt and a base is used in step (i). 5. A process according to any of claims 1 to 4 wherein the compound of formula (I) is reacted with L-ornithine hydrochloride and a base is used in step (i). 6. A process according to any of claims 1 to 5 wherein lithium hydroxide is used as a base in step (i). 7. A process for the preparation of N G -monomethyl-L-arginine hydrochloride comprising: (i) reaction of pyrazole carboxamidine hydrochloride, CH 3 NH NH .HCl with L-ornithine hydrochloride, in a suitable polar solvent, in the presence of lithium hydroxide followed by: (ii) acidification to pH 4 to 5 with hydrochloric acid; then (iii) addition of an alcoholic solvent; then (iv) isolation of N G -monomethyl-L-arginine hydrochloride. 8. A process according to any of claims 1 to 7 whereing the resultant N G - monomethyl-L-arginine hydrochloride is further purified.

Inventors: GILLIES IAIN, GILLIES, IAIN
Applicants: GILLIES IAIN, GLAXO GROUP LTD, GILLIES, IAIN, GLAXO GROUP LIMITED
Application year: 1997
Main classification: C07C277
IPC codes: C07C277, C07C279