Title of Invention

"AN IMPROVED PROCESS FOR THE PREPARATION OF ARTETHERS FROM DIHYDROARTEMISININ"

Abstract

This invention relates to a process for the preparation of artether derivatives of dihydroartemisinin. The dihydroartemisinin is derived from artemisinin, which is a unique sesquiterpene lactone endoperoxide isolated from the plant Artemisia annua L. Artethers prepared for the process of the invention is useful for the treatment of uncomplicated/severe complicated/cerebral and MDR malaria caused mainly by Plasmodius falciparum. The process steps are:dissolving dihydroartemisinin in a mixture of non polar solvent and alcohols in a ratio in the range of 5:1-10:1 under stirring at a temperature in the range of 50-60°C, adding an acid catalyst such as anhydrous aluminum chloride (AICI3) to the reaction mixture and maintaining for a period of 3-4 hours, neutralizing by known method, washing and drying over conventional anhydrous inorganic salts recovering artethers by conventional methods.

Full Text

This invention relates to a process for the preparation of artether derivatives of dihydroartemisinin. The dihydroartemisinin is derived from artemisinin, which is a unique sesquiterpene lactone endoperoxide isolated from the plant Artemisia annua L.~ Artethers prepared for me process of the invention is useful for the treatment of uncomplicated/severe complicated/cerebral and MDR malaria caused mainly by Plasmodiumfalciparum. Approximately, 300-400 million people worldwide now suffer from malaria, and each year 1 -3 million (mostly children) die from this infectious disease. Malaria continues to be one of the most serious public health and environmental problem in a large part of the developing countries including India. Prospects for the control of malaria have been seriously hampered by the emergence of resistance to almost all the available antimalarial drugs such as chloroquine, quinine, primaquine and mefloquine etc. Arteether is a totally new drug introduced in India and has under gone extensive preclinical, animal, toxicological studies as well as clinical studies in Indian subjects as per drug regulatory requirements. Arteether is an ideal antimalarial drug especially for treating drug resistant and complicated P. falcipanim malaria. Arteether shows rapid schizontocidal action with quicker parasite clearance rate and short fever clearance time, virtually with no side effects and low recrudescence rate. Five procedures were reported for the preparation of arteether. First procedure is reported by A. Brossi. B. Young, B. Venugopalan, L. Dominguez Gerpe, H. J. C. Yeh, J. L. Flippen-Anderson. B. Buchs. X. D. Luo. W. Michous and W. Peter (J. Med. Chem., 1988. 31, 646-649). These authors have prepared arteether from dihydroartemisinin of the formula (2) which itself is prepared by sodium borohydride reduction of artemisinin of the formula (1). The dihydroartemisinin of the formula (2) was dissolved in a solvent mixture of ethanol and benzene by heating the solution at 45 °C. After rapid addition of acid catalyst BF3.Et2O, the reaction mixture was refluxed at 70 °C for 1 hr followed by fractional crystallization or chromatography to remove a-isomer. (3-Arteether having formula (3) where R = €2^ (P) was obtained in 60% yield as a crystalline material whereas said a-arteether was obtained as an oil in 19% yield. This method suffers a number of disadvantages. During the process, two isomers, a and P were produced in v; b'-' ratio, which had to be separated by column chromatography, hence additional step is involved leading to over all loss of yield. The a-isomer produced in lesser amount was again equilibrated into 30:70 mixture of a and p-arteether using same experimental procedure, to obtain single isomer. Second method is reported by F. El-Feraly. M. A. Al-Yahya. Kyorabi, D. R. McPhail and A. T. McPhail (J. Nat. Prod., 1992. 55, 878-883) in which anhdrodihydroartemisinin, prepared from artemisinin of the formula (1) was stirred with ethanol in presence ofp-toluenesulfonic acid as a catalyst. Upon work up the product (90%) was found to be a mixture of P-arteether and its C-11 epimer in the ratio of (3:1). In this method only p-arteether is obtained and separation of its C-11 epimer is difficult and preparation of anhydrodihydroartemisinin is a tedious process. This reaction took longer period (22 hr.) to completion. The acid catalyst used in this reaction is required in large amount (60 mg acid catalyst/100 mg dihydroartemisinin). The third method is reported by R. S. Bhakuni, D. C. Jain and R. P. Sharma (Ind. J. Chem., 1995. 3-fB. 529) in which dihydroartemisinin of the formula (3) was stirred with ethanol in benzene at room temperature and chlorotrimethylsilane (CTMS) was used as acid catalyst. After usual work up. a/p-arteether (30:70 mixture of isomers) was obtained in 80-85% yield. The fourth method is reported by A. J. Lin and R. E. Miller (J. Med. Chem. 1995, 38, 764-770) in which the new ether derivatives were prepared by dissolving dihydroartemisinin in anhydrous ether and adding appropriate alcohol followed by boron trifluoride etherate. The reaction mixture was stirred at room temperature for 24 hours. The yield of purified products ranged from 40-90%. Purification was achieved by the use of silica gel chromatography. The fifth method is reported by D. C. Jain, R. S. Bhakuni, S. Saxena, S. Kumar and R. A. Vishwakarma (US Patent 6,346,631) in which ether derivatives of dihydroartemisinin were prepared by dissolving dihydroartemisinin in appropriate dry alcohol followed by cation exchange resin and trialkylorthoformate. The reaction mixture was stirred at room temperature for 10.hours. The yield of the purified products ranged from 80-85%. The above mentioned methods suffer from some disadvantages. Benzene is used as reaction medium, which on work-up left few non-volatile impurities in the reaction product. Also, benzene itself is a potent carcinogenic agent and has been banned in several western countries. All the above-mentioned processes use chemicals, which are not eco-friendly. The minor products formed during the reaction require separation by column chromatography, thereby causing loss of arteether yield. Secondly, the acid catalysts used in the prior arts i.e. chlorotrimethylsilane (CTMS), boron trifluoride etherate (BFa.EtiO) are expensive reagent. It is an object of the invention to provide process for the preparation of artethers by replacing carcinogenic solvent, replacing expensive liquid catalyst used in the prior art by an inexpensive solid acid catalyst thereby making this process a green technology. Accordingly, the present invention provides an improved process for the preparation of artethers (alkyl and benzyl ether derivatives of dihyroartemisinin) from dihydroartemisinin which comprises dissolving dihydroartemisinin in a mixture of non polar solvent and alcohols in a ratio in the range of 5:1-10:1 under stirring at a temperature in the range of 50-60°C, adding an acid catalyst such as anhydrous aluminum chloride (AICI3) to the reaction mixture and maintaining for a period of 3-4 hours, neutralizing by known method, washing and drying over conventional anhydrous inorganic salts recovering artethers by conventional methods. In the improved process of the present invention, the dihydroartemisinin was dissolved in a mixture consisting of non polar solvents such as cyclohexane, n-hexane, toluence and alcohol such as methanol, ethanol, iso-ropanol and benzyl alcohol in the ratio of the range of 10:1-16:1, by stirring at a temperature in the range of 50-600C. Then anhydrous aluminium chloride (AICI3) was added as acid catalyst to the reaction mixture. The ratio of acid catalyst and dihydroartemisinin in the range of 2.25:1-1:1 (w/w). The reaction mixture was left for 3-4 hours at temperature in the range of 50- 60°C. The solvent phase was neutralized by known methods and finally washed with water. Drying the solution over anhydrous inorganic salts such as sodium sulphate, magnesium sulphate. Artether is recovered by conventional methods such as evaporation, chromatography and conventration to yield mixture of α/ ß artether in 84-90% yield. The details of the present invention are provided in the following examples, which are given by way of illustration only and should not be construed to limit the scope of the present invention. Example-1 Step a: Preparation of dihydroartemisinin of the formula (2) from artemisinin of the formula (1) Artemisinin (10 g, 35.46 mmol) in dry methanol (250 ml), was cooled in an ice bath to 0-5 °C for 30 min. To the cooled and stirred solution was added in small portions of NaBKt (4 g, 3 eq.) over a period of 30 minutes. The reaction mixture was stirred at 0-5 °C for 1 hr. after the complete addition of NaBH.4. The crushed ice (800 g) was added to the reaction mixture which produced a white precipitate immediately which was filtered with cold water and dried in a desiccator to obtain pure white powder of dihydroartemisinin (8.9 g, 89% yield) of the formula (2). Step b: Preparation of arteether of the formula (3) where in R = C^H^ Dihydroartemisinin (5.0 g, 17.61 mmol) was dissolved in dry toluene (50 ml) and dry ethanol (5 ml) by stirring the solution for 15 min. at 50-60 °C over an oil bath. Then acid catalyst, anhydrous aluminium chloride (A1C13) (2.35 g, 17.60 mmol, 1 eq.) was added to it and stirring was continued for 3 hours at the same temperature. After completion of the reaction (TLC), the reaction mixture was washed with saturated sodium acetate solution (1 x 50 ml), water (2 x 50 ml). The solution was dried over Na2SC>4 and evaporated. The residue was dissolved in ethyl acetate/w- hexane (4:96) and filtered over a short Sigel column to yield a/p-arteether (4.94 g). The total yield of pure a/p-arteether of the formula (3) where in R = C2H5 was found to be 90% in 29.6:71.8 ratio of its a/p-isomer. The compound is characterized by 'H NMR, IR and mass spectral data and estimated by HPLC. Step c: HPLC analysis of a/p-arteether in reaction mixture HPLC analysis was performed on C-18 column. For estimation, standard solution of detected by UV detector at 215 nm. Chromatograms were plotted and peak areas were calculated. The concentration of a-and p-isomers in samples was determined by using standard curves. Example-2 Preparation of arteether of the formula (3) where in R = CHj Dihydroartemisinin (100 mg, 0.35 mmol) was dissolved in dry toluene (10 ml) and dry methanol (1 ml) by stirring the solution for 15 min. at 50-60 °C over an oil bath. Then acid catalyst, anhydrous aluminium chloride (Aids) (47 mg, 0.35 mmol, 1 eq.) was added to it and stirring was continued for 3 hours at the same temperature. After completion of the reaction (TLC), the reaction mixture was washed with saturated sodium acetate solution (1 x 50 ml), water (2 x 50 ml). The solution was dried over Na?.SO4 and evaporated. The residue was dissolved in ethyl acetate/H-hexane (4:96) and filtered over a short Sigel column to yield a/(3-arteether (3). The total yield of pure a/p-arteether of the formula (3) where in R = CH3 was found to be 85% in 28:72 ratio of its cx/p-isomer. The compound is characterized by *H NMR. IR and mass spectral data. Example-3 Preparation of /so-propyl ether of the formula (3) where in R = Dihydroartemisinin (100 mg, 0.35 mmol) was dissolved in dry toluene (10 ml) and dry iso-propyl alcohol (1 ml) by stirring the solution for 15 min. at 50-60 °C over an oil bath. Then acid catalyst, anhydrous aluminium chloride (A1C13) (47 mg, 0.35 mmol, 1 eq.) was added to it and stirring was continued for 3 hours at the same temperature. After completion of the reaction (TLC), the reaction mixture was worked up as discussed in Example-2. The total yield of pure aJ$-iso-propyl ether of the formula (3) where in R = CH(CH3)2 was found to be 86% in 29.1:71.8 ratio of its a/p-isomer. The compound is characterized by 1H NMR, IR and mass spectral data. ExampIe-4 Preparation of benzyl ether of the formula (3) where in R = Dihydroartemisinin (100 mg, 0.35 mmol) was dissolved in dry toluene (10 ml) and dry benzyl alcohol (1 ml) by stirring the solution for 15 min. at 50-60 °C over an oil bath. Then acid catalyst, anhydrous aluminium chloride (A1C13) (47 mg, 0.35 mmol, 1 eq.) was added to it and stirring was continued for 3 hours at the same temperature. After completion of the reaction (TLC), the reaction mixture was worked up as discussed in Example-2. The total yield of pure a/p-benzyl ether of the formula (3) where in R = CH2C6H5 was found to be 84% in 27:73.2 ratio of its oc/p-isomer. The compound is characterized by 'H NMR, IR and mass spectral data. Example-5 Preparation of butyl ether of the formula (3) where in R = CjHg Dihydroartemisinin (100 mg, 0.35 mmol) was dissolved in dry toluene (10 ml) and dry n-butyl alcohol (1 ml) by stirring the solution for 15 min. at 50-60 °C over an oil bath. Then acid catalyst, anhydrous aluminium chloride (Aids) (47 mg, 0.35 mmol, 1 eq.) was added to it and stirring was continued for 3 hours at the same temperature. After completion of the reaction (TLC), the reaction mixture was worked up as discussed in Example-2. The total yield of pure cc/p-butyl ether of the formula (3) where in R = C^g was found to be 84% in 27:73.2 ratio of its a/p-isomer. The compound is characterized by !H NMR, IR and mass spectral data. Example-6 Dihydroartemisinin was prepared in a similar manner as given in Example-1. Dihydroartemisinin (100 mg, 0.35 mmol) was dissolved in dry toluene (10 ml) and dry ethanol (1 ml) by stirring the solution for 15 min. at 60 °C over an oil bath. Then acid catalyst, anhydrous aluminium chloride (A1C13) (47 mg, 0.35 mmol, 1 eq.) was added to it and stirring was continued for 6 hours at the same temperature. The reaction product showed a number of products with 60% amount of a/p-arteether. Example-7 A solution of dihydroartemisinin (100 mg, 0.35 mmol) in dry toluene (16 ml) and ethanol (1 ml) was treated with the acid catalyst (AlCl?) (47 mg, 0.35 mmol, 1 eq.) and the reaction mixture was left for 20 hr at 30 °C. The reaction mixture furnished 25% yield of a/p-arteether. Example-8 A solution of dihydroartemisinin (100 mg, 0.35 mmol) in dry toluene (10 ml) and ethanol (1 m!) was treated with the acid catalyst (A1C13) (47 mg, 0.35 mmol, 1 eq.) and the reaction mixture was left for 3 hr at 75 °C over an oil bath. The reaction mixture furnished 10% yield of a 'p-arteether and some by-products were also formed. The improved process for the preparation of arteether, the subject matter of this patent, afforded a number of advantages. The main advantages of the process of the present invention are: 1. The overall yield of arteether (oc/p-isomers in 30:70+3% ratio) from artemisinin is 85-90%, which is more than the previously reported methods. 2. Acid catalyst used in our procedure is much cheaper and available as shelf chemical than others methods viz. BP3,/?-TsOH, CTMS, and cation exchange resin. 3. The reaction is carried out at 50-60 °C whereas in the BFs-procedure the reaction is carried out at 75 °C. 4. The ratio of 30:70±3% of a/p-isomer is obtained directly and hence there is no need to make up the ratio, as this ratio of two isomers is more effective in drug development. 5. Crystallization or separation of oc/p-isomers and impurities by chromatography is avoided in the present process. 6. No side products are formed whereas in the BFs-procedure, some side products are formed and in the /7-TsOH procedure an unwanted C-l 1 epimer is formed. 7. This process is a general method for the preparation of different ether derivatives of dihydroartemisinin. We claim 1. An improved process for the preparation of artethers (alkyl and benzyl ether derivatives of dihyroartemisinin) from dihydroartemisinin which comprises dissolving dihydroartemisinin in a mixture of non polar solvent and alcohols in a ratio in the range of 5:1-10:1 under stirring at a temperature in the range of 50-60°C, adding an acid catalyst such as anhydrous aluminum chloride (AICI3) to the reaction mixture and maintaining for a period of 3-4 hours, neutralizing by known method, washing and drying over conventional anhydrous inorganic salts recovering artethers by conventional methods. 2. An improved process as claimed in claim 1, wherein the non-polar solvent used is selected from cyclohexane , n-hexane and toluene. 3. An improved process as claimed in claim 1, wherein alcohol used is selected from methanol, ethanol iso-propanol butanol and benzy alcohol. 4. An improved process as claimed in claim 1, where dihydroartemisinin and alcohol is in the ratio 1:1-0.1:1 (w/v). 5. An improved process as claimed in claim 1, wherein dihydroartemisinin and acid catalyst is in the ratio 2.25:1-1:1 (w/w). 6. An improved process as claimed in claim 1, wherein recovery of artethers is affected by conventional methods such as evaporation, chromatography and concentration. 7. An improved process as claimed in claim 1, wherein the overall yield of arteether (a/ p -isomers in 30:70+3% ratio) from artemisinin is 85-90 8. An improved process for the preparation of artethers from dihydroartemisinin substantially as herein described with reference to the examples and drawing accompanying this specifications.

Documents:

542-DEL-2003-Abstract-(27-02-2009).pdf

542-del-2003-abstract.pdf

542-DEL-2003-Claims-(27-02-2009).pdf

542-del-2003-claims.pdf

542-DEL-2003-Correspondence-Others-(27-02-2009).pdf

542-del-2003-correspondence-others.pdf

542-del-2003-correspondence-po.pdf

542-DEL-2003-Description (Complete)-(27-02-2009).pdf

542-del-2003-description (complete).pdf

542-del-2003-drawings.pdf

542-del-2003-form-1.pdf

542-del-2003-form-18.pdf

542-del-2003-form-2.pdf

542-DEL-2003-Form-3-(27-02-2009).pdf