WO2012131541A1 - A process for the isolation of emtricitabine - Google Patents
A process for the isolation of emtricitabine Download PDFInfo
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- WO2012131541A1 WO2012131541A1 PCT/IB2012/051354 IB2012051354W WO2012131541A1 WO 2012131541 A1 WO2012131541 A1 WO 2012131541A1 IB 2012051354 W IB2012051354 W IB 2012051354W WO 2012131541 A1 WO2012131541 A1 WO 2012131541A1
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D411/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen and sulfur atoms as the only ring hetero atoms
- C07D411/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen and sulfur atoms as the only ring hetero atoms containing two hetero rings
- C07D411/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen and sulfur atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring-member bond
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- the present invention relates to a process for the preparation of 4-amino-5-fluoro-l- [(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone, hereinafter referred to as emtricitabine, represented by the compound of formula I. More particularly, the present invention relates to a simplified process for the isolation of emtricitabine from a reaction mixture.
- emtricitabine 4-amino-5-fluoro-l-[(2R,5S)-2- (hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone and it is structurally represented herein below as formula I
- Emtricitabine is an analogue of cytidine and as illustrated in formula I, is the cis- enantiomer having 2R,5S absolute configuration. It is a nucleoside reverse transcriptase inhibitor used for the treatment of Human Immunodeficiency Virus (HIV) infections in adults and children, which works by inhibiting reverse transcriptase, the enzyme that copies HIV RNA into new viral DNA.
- Emtricitabine is available in the market under the trade name Emtriva® (formerly known as Coviracil®).
- Emtricitabine is also available in the market under the trade name Truvada®, as a fixed dose in combination with tenofovir.
- Emtriva® previously known as Coviracil®
- Truvada® Truvada®
- the process involves enzymatic or stereoselective chemical synthesis for the preparation of chiral emtricitabine.
- the stereoselective chemical synthesis particularly comprises reaction of (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate, hereinafter referred to as 'FCME' and represented by the compound of formula II, with a reducing agent in the presence of a solvent to obtain a salt of emtricitabine and subsequent conversion of the salt of emtricitabine to obtain emtricitabine free base, involving tedious and lengthy methods of separation, since the product emtricitabine is highly soluble in polar media, difficulties are encountered in the process of isolation of emtricitabine from such media.
- 5,696,254 discloses a stereoselective chemical synthesis for the preparation of emtricitabine of formula I.
- the process comprises steps of: (i) reaction of FCME of formula II with lithium aluminum hydride (LAH) as a reducing agent in the presence of tetrahydrofuran (THF) as a solvent at an ambient temperature under an argon atmosphere to obtain a reaction mixture, (ii) the reaction mixture was stirred for 2 hours and was quenched by addition of methanol followed by silica gel, (iii) the reaction mixture was then passed through a short pad of celite and silica gel and eluted with a 1: 1: 1 mixture of ethyl acetate (EtOAc)-hexane-methanol, (iv) the elute was concentrated and subjected to column chromatography separation to yield a gummy solid, (v) the gummy solid was then dried azeotropically with toluene to obtain emtric
- US Patent No. 6,051,709 discloses a process for the preparation of both lamivudine and emtricitabine, however, a particular example describing the process for the preparation of lamivudine is provided in the US '709 Patent.
- the structural difference between the lamivudine and emtricitabine is such that, in lamivudine the pyrimidine moiety is 'cytosine', whereas in emtricitabine the pyrimidine moiety is '5- fluorocytosine' .
- lamivudine is a non-fluoro analogue of emtricitabine of formula I.
- the US '709 Patent involves isolation of monosalicylate salt of lamivudine from the reaction mixture and since lamivudine is highly soluble in polar solvent, isolation of lamivudine from the reaction mixture becomes very difficult, whereas the salts of lamivudine are efficiently isolated from the polar solvent as having poor aqueous solubility.
- US Patent No. 7,534,885 particularly describes a process for the salification of FCME of formula II to obtain a salt of FCME and subsequent conversion of the salt of FCME to obtain emtricitabine formula I.
- the US'885 Patent has mainly discussed a process for the preparation of FCME oxalate.
- Said US Patent further provides a process for the conversion of FCME oxalate to emtricitabine comprising the steps of: (i) reaction of FCME oxalate with a solution of sodium borohydride and 30% sodium hydroxide in water in the presence of a mixture of tetrahydrofuran, methanol and water as a solvent, potassium carbonate and dipotassium hydrogen phosphate, (ii) the reaction mixture obtained was then stirred at a temperature of 20°C-25°C for 1 hour, (iii) then the pH of the reaction mixture was adjusted to 4 using 37% hydrochloric acid and the organic solvents were evaporated off under vacuum, (iv) the remaining aqueous phase solution was extracted three times with toluene and the pH of the aqueous solution again adjusted to 7.3-7.4 using 30% sodium hydroxide solution and water was evaporated off under vacuum to obtain a residue, (v) the residue was then taken up in isopropanol and evaporated under vacuum, (vi) the
- the US'885 Patent utilizes salt of FCME, such as oxalate salt of FCME for the preparation of emtricitabine. Moreover, after the reaction completion, pH of the whole reaction mixture was adjusted to 4 using 37% hydrochloric acid and then the organic solvents were evaporated off under vacuum, this leads to the handling of large volume of reaction mixture. Also the step (viii) of the above discussed process requires overnight stirring for separation of the product, emtricitabine with a yield of only 31%. Therefore, the process of the US'885 Patent, as discussed herein above for the preparation of emtricitabine is disadvantageous for the industrial manufacturing of emtricitabine.
- US Patent Application Publication No. 2010/0256372 discloses a process for the preparation of emtricitabine of formula I comprising the steps of: (i) reducing the compound FCME of formula II employing the reducing agent in the presence of organic solvents to obtain reaction mixture containing emtricitabine, (ii) followed by insitu salification of emtricitabine using an organic acid or a mineral acid in the presence of an organic solvent to yield organic or mineral acid salt of emtricitabine, such as hydrochloride, hydrobromide or methane sulfonate salt of emtricitabine, (iii) neutralizing the product obtained in step (ii) using an organic amine bases in the presence of organic solvent to yield emtricitabine.
- the process disclosed in the US '372 patent appln. involves three steps process for the preparation of emtricitabine, involving first step of reduction of FCME to emtricitabine, second step of isolation of salt of emtricitabine from the reaction mixture and third step of conversion of salt of emtricitabine to emtricitabine free base, thereby making the process lengthy and costly. Also the process involves lengthy work-up process for the isolation of salt of emtricitabine as well as emtricitabine free base from the reaction mixture, thus renders the process for preparation of emtricitabine industrially not viable.
- the process for the preparation of emtricitabine of formula I can be improved particularly in terms of industrial applicability, by providing cost-effective and simple isolation process for emtricitabine from the reaction mixture, that would result in obtaining emtricitabine in good yield and having high purity.
- the process for the isolation of emtricitabine disclosed in the cited prior art references mostly suggests insitu salification of emtricitabine for the isolation of the salt of emtricitabine such as monosalicylate, oxalate, hydrochloride, hydrobromide, methane sulfonate etc.
- emtricitabine of formula I can be obtained in good yield and high purity from (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II through an improved process, which although involves use of sodium borohydride as a reducing agent in the presence of phosphate buffer, provides a single step process for the preparation of emtricitabine from FCME and avoids preparation of any salt of emtricitabine as an intermediate step, which in turn reduces the number of steps.
- the present invention provides a simple, cost-effective, industrially applicable process for the isolation of emtricitabine, which is used as a nucleoside reverse transcriptase inhibitor.
- An object of the present invention is to provide a process for the isolation of emtricitabine of formula I from a reaction mixture obtained by reaction of the compound of formula II with a reducing agent in the presence of phosphate buffer and a solvent, resulting in obtaining emtricitabine having yield of > 60% and purity of > 99.8%.
- Another object of the present invention is to provide an alternate process for the isolation of emtricitabine of formula I from a reaction mixture obtained by reaction of the compound of formula II with a reducing agent in the presence of phosphate buffer and a solvent, resulting in obtaining emtricitabine having purity > 99.8%.
- Yet another object of the present invention is to provide a process for the isolation of emtricitabine of formula I, which is simple and cost-effective, and avoids isolation of salts of emtricitabine.
- Still another object of the present invention is to provide a process for the isolation of emtricitabine of formula I, which is industrially applicable.
- step (c) adding an aqueous acid to the mixture of step (b) to adjust pH of the reaction mixture to 8.0 to 8.5, d) distilling out the polar solvent from the mixture of step (c) to obtain an aqueous solution,
- step (d) extracting the aqueous solution of step (d) with a non-polar solvent and distilling out the aqueous solution to obtain a liquid
- step (e) diluting the liquid of step (e) with a polar solvent and distilling out the solvent to remove the traces of water from the reaction mixture
- step (f) diluting the mixture of step (f) with a polar solvent and refluxing the reaction mixture at a temperature of 85°C to 90°C for 30 minutes to precipitate out an inorganic residue
- step (g) filtering off the inorganic residue obtain in step (g) and distilling out the filtrate to obtain a residue
- step (h) diluting the residue of step (h) with an alcohol and heating it for 30 minutes at a temperature of 85°C to 90°C, and
- step (i) cooling the reaction mixture of step (i) at a temperature of 10°C to 15°C and isolating the precipitated emtricitabine by filtration, having purity of >99.8 .
- step (iii) adding an aqueous acid to the mixture of step (ii) to adjust pH of the reaction mixture to 8.0 to 8.5,
- step (iv) distilling out the polar solvent from the mixture of step (iii) to obtain an aqueous solution
- step (v) extracting the aqueous solution of step (iv) with a non-polar solvent
- step (vi) diluting the aqueous solution of step (v) with polar solvent at a temperature ranging from 15°C to 35°C, and
- step (vii) cooling the reaction mixture of step (vi) slowly at a temperature of 5°C to 10°C over a period of 6 to 12 hours and isolating the precipitated emtricitabine by filtration, having purity of > 99.8%.
- the process for isolation of emtricitabine of formula I overcomes the disadvantages associated with the processes disclosed in the cited prior art, which mainly concerns with the isolation of salt of emtricitabine from the reaction mixture.
- the reason being emtricitabine is highly soluble in the solvents used during the reaction and thereby facing the difficulties to isolate pure emtricitabine free base from the reaction mixture.
- the present invention provides a solution to this problem by providing the processes that involve use of non-hazardous and inexpensive reducing agents such as sodium borohydride, thereby avoiding use of hazardous reducing agents such as lithium aluminum hydride (LAH) as discussed in the prior art.
- LAH lithium aluminum hydride
- the process for the isolation of emtricitabine of formula I from the reaction mixture provides the desired emtricitabine free base with good yield and high purity, without use of any lengthy isolation process and hazardous reducing agents, thereby making the process of the isolation of emtricitabine simple, cost-effective and industrially applicable.
- the present invention relates to a process for the isolation of 4-amino-5-fluoro-l- [(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture comprising the steps of:
- step (b) adding an aqueous acid to the mixture of step (b) to adjust pH of the reaction mixture to 8.0 to 8.5
- step (c) distilling out the polar solvent from the mixture of step (c) to obtain an aqueous solution
- step (d) extracting the aqueous solution of step (d) with a non-polar solvent and distilling out the aqueous solution to obtain a liquid
- step (e) diluting the liquid of step (e) with a polar solvent and distilling out the solvent to remove the traces of water from the reaction mixture
- step (f) diluting the mixture of step (f) with a polar solvent and refluxing the reaction mixture at a temperature of 85°C to 90°C for 30 minutes to precipitate out an inorganic residue
- step (g) filtering off the inorganic residue obtain in step (g) and distilling out the filtrate to obtain a residue
- step (h) diluting the residue of step (h) with an alcohol and heating it for 30 minutes at a temperature of 85°C to 90°C, and
- step (i) cooling the reaction mixture of step (i) at a temperature of 10°C to 15°C and isolating the precipitated emtricitabine by filtration, having purity of >99.8 .
- step (a) of the process said FCME of formula II is reacted with sodium borohydride as a reducing agent in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent to obtain the reaction mixture.
- the reaction is carried out in the presence of a polar solvent selected from alcohols, ethers or a mixture thereof.
- a polar solvent selected from alcohols, ethers or a mixture thereof.
- the ether used for the reaction as a solvent is selected from tetrahydrofuran, dioxane, diethyl ether or dimethyl ether or a mixture thereof.
- the alcohol used for the reaction as a solvent is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof, preferably the solvent used for the reaction is isopropyl alcohol.
- the process involves: taking isopropyl alcohol in a reaction vessel and a solution of dipotassium hydrogen phosphate in water is charged.
- the solution of sodium borohydride in aqueous sodium hydroxide is added slowly to the reaction mixture over a period of 1 to 1.5 hours. It has been indicated herein above that the inventors of the present invention performed numerous experiments to develop an efficient process for the isolation of emtricitabine of formula I from the reaction mixture obtained by reacting FCME of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to 20°C.
- the process pertaining to step (a) of the present invention was carried out at varying temperature conditions with the constant volume of a polar solvent. The results of effect of varying temperature on the time required for the completion of reaction and the purity of the emtricitabine obtained are presented in the following Table- 1.
- step (b) of the process the aqueous and organic layers formed in the reaction mixture are separated using a separation flask.
- the organic layer containing the polar solvent such as isopropyl alcohol is used as such for the next step.
- the pH of the organic layer is approximately 11.2.
- the reaction mixture obtained in step (b) is adjusted to pH of 8.0 to 8.5 using an aqueous acid.
- the aqueous acid used in the reaction is a mineral acid selected from hydrochloric acid and hydrobromic acid, or a weak organic acid selected from formic acid, acetic acid or oxalic acid or a mixture thereof, preferably hydrochloric acid is used as an aqueous acid in the reaction.
- the hydrochloric acid used in the reaction to adjust the pH of the reaction mixture to 8.0 to 8.5 is 20% (v/v) dilute hydrochloric acid.
- step (d) of the process in the step (d) of the process, the reaction mixture obtained in step (c) is distilled out at a temperature of 40°C to 45°C under vacuum to obtain an aqueous solution.
- the aqueous solution obtained in step (d) is extracted with a non-polar solvent.
- the non-polar solvent used for the step (e) of the process is selected from hexane, cyclohexane, toluene, benzene, dioxane, chloroform or diethyl ether or mixture thereof, preferably the non-polar solvent used for the reaction is toluene.
- the aqueous solution obtained in step (e) is extracted with a non-polar solvent to remove the byproduct menthol from the reaction mixture, which is formed during the reaction.
- the aqueous solution obtained after the extraction with non-polar solvent such as toluene may optionally be treated with charcoal for 30 minutes at a temperature of 25°C to 30°C and the reaction mixture obtained is filtered off. The filtrate obtained is distilled out under vacuum at 50°C to 60°C to obtain a liquid.
- the liquid obtained in step (e) is diluted with a polar solvent such as isopropyl alcohol and distill out the polar solvent at a temperature of 50°C to 60°C under vacuum.
- the liquid obtained in step (e) is treated with a polar solvent for the complete removal of water content from the reaction mixture.
- a polar solvent for the complete removal of water content from the reaction mixture.
- the presence of water in the reaction mixture retains the water soluble inorganic residue in the reaction mixture, which will get extracted along with the product in the subsequent steps.
- the product, emtricitabine of formula I may fail in the sulfated ash test.
- emtricitabine has relatively good solubility in water, the presence of traces water in the reaction mixture may also result in loss of yield of the product.
- the reaction mixture may be treated with a polar solvent for multiple numbers of times.
- the polar solvent obtained is distilled out from the reaction mixture at a temperature of 50°C to 60°C under vacuum to obtain a thick solid.
- the thick solid obtained is analyzed for moisture content and the limit for moisture content is not more than (NMT) 1.5%.
- the thick solid obtained in step (f) is diluted with a polar solvent at a temperature of 50°C to 60°C and reflux the reaction mixture at a temperature of 85°C to 90°C for 30 minutes, the product, emtricitabine of formula I gets extracted into the polar solvent and the undissolved inorganic residue precipitated out from the reaction mixture.
- step (h) of the process the reaction mixture obtained in step (g) is cooled and filtered to remove undissolved inorganic residue from the reaction mixture.
- the filtrate obtain is then distilled out at a temperature of 45°C to 50°C under vacuum to obtain a residue.
- the residue obtained in step (h) may diluted with alcohol and distilled out alcohol at a temperature of 45°C to 50°C under vacuum to obtain the residue.
- the solvent of the reaction mixture is changed from isopropyl alcohol to ethanol.
- the alcohol used in the reaction is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof, preferably said alcohol is ethanol.
- the residue obtained at stage (h) is diluted with 2.5 volume of alcohol and the reaction mixture is heated at a temperature of 85°C to 90°C for 30 minutes.
- step (j) of the process in the step (j) of the process, the reaction mixture obtained in step (i) is cooled to a temperature of 10°C to 15°C and the reaction mixture is stirred at a temperature of 10°C to 15°C for 4 hours to precipitate the product, emtricitabine of formula I.
- the product obtained under vacuum is filtered and washed with ethanol.
- the product is dried under vacuum at a temperature of 45°C to 50°C.
- the emtricitabine obtained by using the above discussed process having yield > 60% and purity > 99.8%.
- the present invention also relates to an alternate process for the isolation of 4-amino- 5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture comprising the steps of:
- step (iii) adding an aqueous acid to the mixture of step (ii) to adjust pH of the reaction mixture to 8.0 to 8.5,
- step (iv) distilling out the polar solvent from the mixture of step (iii) to obtain an aqueous solution
- step (v) extracting the aqueous solution of step (iv) with a non-polar solvent
- step (vi) diluting the aqueous solution of step (v) with polar solvent at a temperature ranging from 15°C to 35°C, and
- step (vii) cooling the reaction mixture of step (vi) slowly at a temperature of 5°C to 10°C over a period of 6 to 12 hours and isolating the precipitated emtricitabine by filtration having purity of > 99.8%.
- step (i) of the process said FCME of formula II is reacted with sodium borohydride as a reducing agent in the presence of dipotassium hydrogen phosphate and 9 volume of polar solvent to obtain a reaction mixture.
- the reaction is carried out in the presence of a polar solvent selected from alcohols, ethers or a mixture thereof.
- a polar solvent selected from alcohols, ethers or a mixture thereof.
- the ether used for the reaction as a solvent is selected from tetrahydrofuran, dioxane, diethyl ether or dimethyl ether or a mixture thereof.
- the alcohol used for the reaction as a solvent is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof, preferably the solvent used for the reaction is isopropyl alcohol.
- the process involves: taking isopropyl alcohol in a reaction vessel and a solution of dipotassium hydrogen phosphate in water is charged.
- step (ii) of the process the aqueous and organic layers formed in the reaction mixture are separated using a separation flask.
- the organic layer containing the polar solvent such as isopropyl alcohol is used as such for the next step.
- the pH of the organic layer is approximately 11.2.
- the reaction mixture obtained in step (ii) of the process is adjusted to pH of 8.0 to 8.5 using an aqueous acid.
- the aqueous acid used in the reaction is a mineral acid selected from hydrochloric acid and hydrobromic acid, or a weak organic acid selected from formic acid, acetic acid or oxalic acid or a mixture thereof, preferably hydrochloric acid is used as an aqueous acid in the reaction.
- the hydrochloric acid used in the reaction to adjust the pH of the reaction mixture to 8.0 to 8.5 is 20% (v/v) dilute hydrochloric acid.
- the reaction mixture obtained in step (iii) is distilled out at a temperature of 40°C to 45°C under vacuum to obtain an aqueous solution.
- the aqueous solution obtained in step (iv) is extracted with a non-polar solvent.
- the non-polar solvent used for the step (v) is selected from hexane, cyclohexane, toluene, benzene, dioxane, chloroform or diethyl ether or mixture thereof, preferably the non-polar solvent used for the reaction is toluene.
- the aqueous solution obtained in step (iv) is extracted with a non-polar solvent to remove the by- product menthol from the reaction mixture, which is formed during the reaction.
- the aqueous solution obtained in step (v) may optionally stirred at a temperature of 15°C to 35°C for 15 to 20 minutes.
- the aqueous solution is then diluted with 2 volumes of polar solvent, such as isopropyl alcohol at a temperature of 15°C to 35°C.
- the reaction mixture in the step (vii) of the process, slowly cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours to precipitate the product, emtricitabine of formula I.
- the precipitated product is then filtered and washed with isopropyl alcohol.
- the product dried under vacuum at a temperature of 45°C to 50°C, having yield > 50% and purity > 99.8%.
- step (vi) and step (vii) of the present invention performed numerous experiments to develop an efficient process for the isolation of emtricitabine of formula I.
- the process discussed herein above pertaining to step (vi) and step (vii) of the present invention was carried out under different reaction conditions particularly, the results of effect of varying volumes of solvent at different temperature conditions and at varying time duration on the yield of emtricitabine are presented in the following Table-3.
- step (v) when the aqueous solution obtained in step (v) is diluted with 1 volume of a polar solvent, based on the compound of formula II, at a temperature of 15°C to 35°C and then the reaction mixture is slowly cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours, it results with 30% yield of emtricitabine of formula I.
- step (v) when the aqueous solution obtained in step (v) is diluted with 2 volume of polar solvent, based on the compound of formula II, at a temperature of 15°C to 35°C and then the reaction mixture is slowly cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours, it results with increase in the yield of emtricitabine of formula I, i.e. emtricitabine is obtained in a yield of 50% (with respect to Ex. No. 1).
- reaction mixture Further diluted the reaction mixture with 3 rd lot of polar solvent and reflux the reaction mixture to a temperature of 85°C to 90°C for 30 minutes to obtain the residue. Cool the reaction mixture and filter, the filtrate obtained is then distilled out at a temperature of 45°C to 50°C to obtain the residue. The residue obtained is then diluted with alcohol such as ethanol and distilled out the alcohol at a temperature of 45°C to 50°C. The reaction mixture further diluted with 2 nd lot of alcohol and the reaction mixture obtained is heated at a temperature of 85°C to 90°C for 30 minutes and cool the reaction mixture at a temperature of 10°C to 15°C to obtain the product, emtricitabine. Filter the product obtained and wash with ethanol. Dry the product under vacuum.
- alcohol such as ethanol
- the emtricitabine obtained from the above discussed process having yield > 60% and purity > 99.8%.
- the emtricitabine of formula I is isolated from the reaction mixture according to the alternate process described herein below: To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, charged dipotassium hydrogen phosphate and water and stirred the reaction mixture for 15 minutes. Then charged polar solvent, such as isopropyl alcohol to the reaction vessel and further stirred the reaction mixture for 15 minutes and the reaction mixture obtained is cooled to a temperature of 15°C to 20°C.
- polar solvent such as isopropyl alcohol
- the aqueous solution obtained is extracted with non-polar solvent such as toluene
- the aqueous layer obtained is diluted with a polar solvent at a temperature of 15°C to 20°C and slowly cooled to a temperature of 5°C to 10°C over a period of 6-12 hours to obtain the product, emtricitabine. Filter the product obtained and wash with polar solvent.
- the emtricitabine obtained from the above discussed process having yield > 50% and purity > 99.8%.
- FCME cyclopentamino-5- fluoro-2-oxo-l(2H)-pyrimidinyl)-l,3-oxathiolane-2-carboxylate
- the process involves reaction of 5-fluorocytosine with 2,,4,6-collidine and t-butyldimethylsilyl trifluoromethane sulfonate in the presence of dichloromethane, the resultant reaction mixture was stirred for 15 minutes to obtain the clear solution.
- a separately prepared solution of (lR,2S,5R)-menthyl-5R-acetoxy-l,3-oxathiolane-2S-carboxylate in dichloromethane was added to the clear solution, followed by iodotrimethylsilane and stirred for 3 hours. Further reaction work-up yields the product FCME of formula II.
- reaction mixture further stirred for 4 - 5 hours and the two layers formed were separated.
- 20% (v/v) dilute hydrochloric acid (4.6ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was further stirred for 15 minutes.
- the reaction mixture was then distilled out and the aqueous solution obtained was extracted with toluene (200ml).
- the reaction mixture then treated with charcoal (5g) and filtered.
- the filtrate obtained was distilled out to obtain a liquid and diluted with isopropyl alcohol (200ml), distilled out the isopropyl alcohol. Diluted the reaction mixture again with isopropyl alcohol (200ml) and distilled out.
- the reaction mixture further diluted with isopropyl alcohol (400ml) and refluxed the reaction mixture to a temperature of 85°C to 90°C for 30 minutes to obtain the residue. Cooled the reaction mixture and filtered, the filtrate obtained was then distilled out to obtain the residue. Diluted the residue obtained with ethanol (100ml) and distilled out.
- the reaction mixture again diluted with ethanol (200ml) and heated at a temperature of 85°C to 90°C for 30 minutes and then cooled at a temperature of 10°C to 15°C to precipitate the product, emtricitabine. The resulting product was filtered and washed with ethanol. Dry the product under vacuum.
- Example 2 To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, dipotassium hydrogen phosphate (73.8g) and water (165ml) was charged and stirred the reaction mixture for 15 minutes. To the reaction mixture then charged isopropyl alcohol (350ml) and further stirred for 15 minutes and cooled the reaction mixture to a temperature of 15°C to 20°C. Then charged FCME (50g) dissolved in isopropyl alcohol (100ml) to the reaction mixture and stirred at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride (13g) dissolved in aqueous sodium hydroxide was added dropwise to the reaction mixture over the period of 1-1.5 hours.
- reaction mixture further stirred for 4 - 5 hr and the two layers formed were separated.
- 20% (v/v) dilute hydrochloric acid (1.8ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was further stirred for 15 minutes.
- the reaction mixture was distilled out and the aqueous solution obtained was extracted with toluene (100ml).
- the reaction mixture then treated with charcoal (2.5g) and filtered.
- the filtrate obtained was distilled out to obtain a liquid and diluted with isopropyl alcohol (50ml), distilled out the isopropyl alcohol.
- the reaction mixture was again diluted with isopropyl alcohol (100ml) and distilled out.
- reaction mixture further stirred for 4 - 5 hours and the two layers formed were separated.
- 20% (v/v) dilute hydrochloric acid (4ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was stirred for 15 minutes.
- the reaction mixture was distilled out and the aqueous solution obtained was extracted with toluene (200ml).
- the aqueous solution obtained was then diluted with isopropyl alcohol (200ml) at a temperature of 15°C to 20°C and slowly cooled to a temperature of 5°C to 10°C over a period of 6- 12 hours to obtain the product, emtricitabine.
- the product was filtered and washed with isopropyl alcohol.
- reaction mixture further stirred for 4 - 5 hours and the two layers formed were separated.
- 20% (v/v) dilute hydrochloric acid (2ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was stirred for 15 minutes.
- the reaction mixture was distilled out and the aqueous solution obtained was extracted with toluene (100ml).
- the aqueous solution obtained was then diluted with isopropyl alcohol (100ml) at a temperature of 15°C to 20°C and slowly cooled to a temperature of 5°C to 10°C over a period of 6- 12 hours to obtain the product, emtricitabine.
- the product was filtered and washed with isopropyl alcohol.
- Buffer preparation 2.72g/L of monobasic potassium phosphate and 4.32g/L of 1- octanesulfonic acid sodium salt in water. Adjust with 20% phosphoric acid to a pH of 2.2.
Abstract
The present invention provides a process for the isolation of 4-amino-5-fluoro-1- [(2R,5S)-2-(hydroxymethyl)-1,3-oxathiolan-5-yl]-2(1H)-pyrimidinone (emtricitabine) of formula I from the reaction mixture obtained by reacting (1R,2S,5R)-menthyl- (2S,5R)-5-(4-amino-5-fluoro-2-oxo-1(2H)-pyrimidinyl)-1,3-oxathiolane-2- carboxylate (FCME) of the formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent at a temperature of 15°C-20°C, avoiding isolation of salt of emtricitabine, such as oxalate, monosalicylate, hydrochloride, hydrobromide, methane sulfonate etc. and subsequent convention of salt of emtricitabine to emtricitabine free base; using simple, cost-effective and industrially applicable process.
Description
A PROCESS FOR THE ISOLATION OF EMTRICITABINE
FIELD OF THE INVENTION
The present invention relates to a process for the preparation of 4-amino-5-fluoro-l- [(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone, hereinafter referred to as emtricitabine, represented by the compound of formula I. More particularly, the present invention relates to a simplified process for the isolation of emtricitabine from a reaction mixture.
BACKGROUND OF THE INVENTION
The chemical name of emtricitabine is 4-amino-5-fluoro-l-[(2R,5S)-2- (hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone and it is structurally represented herein below as formula I
Formula I Emtricitabine is an analogue of cytidine and as illustrated in formula I, is the cis- enantiomer having 2R,5S absolute configuration. It is a nucleoside reverse transcriptase inhibitor used for the treatment of Human Immunodeficiency Virus (HIV) infections in adults and children, which works by inhibiting reverse transcriptase, the enzyme that copies HIV RNA into new viral DNA. Emtricitabine is available in the market under the trade name Emtriva® (formerly known as Coviracil®). Emtricitabine is also available in the market under the trade name Truvada®, as a fixed dose in combination with tenofovir.
The process for the preparation of emtricitabine of formula I is disclosed in several prior art documents. Generally, the process involves enzymatic or stereoselective chemical synthesis for the preparation of chiral emtricitabine. The stereoselective chemical synthesis particularly comprises reaction of (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate, hereinafter referred to as 'FCME' and represented by the compound of formula II, with a reducing agent in the presence of a solvent to obtain a salt of emtricitabine and subsequent conversion of the salt of emtricitabine to obtain emtricitabine free base, involving tedious and lengthy methods of separation, since the product emtricitabine is highly soluble in polar media, difficulties are encountered in the process of isolation of emtricitabine from such media.
The product, emtricitabine, (-)-enantiomer of 4-amino-5-fluoro-l-[(2R,5S)-2- (hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone of formula I, which is at least 95% free of the corresponding (+)-enantiomer is disclosed in the US Patent No. 6,703,396. The patent further discloses a process for the preparation of (-)-enantiomer of emtricitabine involving an enzymatic process for the resolution of racemic mixture of emtricitabine. US Patent No. 5,696,254 discloses a stereoselective chemical synthesis for the preparation of emtricitabine of formula I. The process comprises steps of: (i) reaction of FCME of formula II with lithium aluminum hydride (LAH) as a reducing agent in the presence of tetrahydrofuran (THF) as a solvent at an ambient temperature under an argon atmosphere to obtain a reaction mixture, (ii) the reaction mixture was stirred for 2 hours and was quenched by addition of methanol followed by silica gel, (iii) the reaction mixture was then passed through a short pad of celite and silica gel and eluted with a 1: 1: 1 mixture of ethyl acetate (EtOAc)-hexane-methanol, (iv) the elute was concentrated and subjected to column chromatography separation to yield a gummy solid, (v) the gummy solid was then dried azeotropically with toluene to obtain emtricitabine in 80 % yield. Even though the process disclosed in the US'254 Patent provides emtricitabine with 80% yield, the process involves use of hazardous reagents and tedious process for the isolation of emtricitabine. The process involves use of LAH as a reducing agent, which is highly corrosive in nature. LAH on contact
with moisture forms lithium hydroxide, which can cause severe burns. Also powdered LAH forms dusts that can pose an inhalation hazard. Moreover, the product emtricitabine isolated from the reaction mixture using column chromatography is obtained as a gummy solid, which on further azeotrophic distillation yields emticitabine, thereby making the above discussed process of the US'254 Patent tedious, hazardous and industrially disadvantageous.
US Patent No. 6,051,709 discloses a process for the preparation of both lamivudine and emtricitabine, however, a particular example describing the process for the preparation of lamivudine is provided in the US '709 Patent. The structural difference between the lamivudine and emtricitabine is such that, in lamivudine the pyrimidine moiety is 'cytosine', whereas in emtricitabine the pyrimidine moiety is '5- fluorocytosine' . Thus, lamivudine is a non-fluoro analogue of emtricitabine of formula I. The process disclosed in said US Patent for the preparation of lamivudine comprising the steps of: (i) a process for the preparation of (2R,5R)-5-hydroxy- [l,3]oxathiolane-2-carboxylic acid, 2S-isopropyl-5R-methyl-lR-cyclohexyl ester from L-menthyl glyoxylate hydrate and dithianediol, (ii) a process for the preparation of (2R,5R)-5-(4-amino-2-oxo-2H-pyrimidin-l-yl)-[l,3]oxathiolane-2-carboxylic acid, 2S-isopropyl-5R-methyl-lR-cyclohexyl ester from the product obtained in step (i), (iii) a process for the preparation of 4-amino-l-(2R-hydroxymethyl-[l,3]oxathiolan- 5S-yl)-lH-pyrimidin-2-one monosalicylate, which involves reaction of the product obtained in step (ii) with a solution of sodium borohydride in water containing sodium hydroxide solution in the presence of dipotassium hydrogen phosphate and industrial methylated spirit (IMS) as a solvent, followed by reaction with salicylic acid to yield lamivudine monosalicylate, (iv) the resulting lamivudine monosalicylate obtained in step (iii) on treatment with triethylamine in the presence of IMS as a solvent yields lamivudine. The US '709 Patent involves isolation of monosalicylate salt of lamivudine from the reaction mixture and since lamivudine is highly soluble in polar solvent, isolation of lamivudine from the reaction mixture becomes very difficult, whereas the salts of lamivudine are efficiently isolated from the polar solvent as having poor aqueous solubility. The process for preparation of lamivudine or its analogue emtricitabine described in said patent necessitates isolation of salt of lamivudine or its analogue and then conversion of the salt of lamivudine or its
analogue to free base of lamivudine or its analogue, thereby making the process lengthy and tedious and hence, not viable for industrial manufacturing. Moreover, although the process disclosed in the US'709 Patent appears to be applicable for lamivudine, without any implementation difficulties, the process is practically inapplicable when utilized for the preparation of emtricitabine.
The process disclosed in the US Patent No. 6,051,709 is schematically depicted herein below:
Lamivudine
US Patent No. 7,534,885 particularly describes a process for the salification of FCME of formula II to obtain a salt of FCME and subsequent conversion of the salt of FCME to obtain emtricitabine formula I. The US'885 Patent has mainly discussed a process for the preparation of FCME oxalate. Said US Patent further provides a process for the conversion of FCME oxalate to emtricitabine comprising the steps of: (i) reaction of FCME oxalate with a solution of sodium borohydride and 30% sodium hydroxide in water in the presence of a mixture of tetrahydrofuran, methanol and water as a solvent, potassium carbonate and dipotassium hydrogen phosphate, (ii) the reaction mixture obtained was then stirred at a temperature of 20°C-25°C for 1 hour, (iii) then the pH of the reaction mixture was adjusted to 4 using 37% hydrochloric acid and the organic solvents were evaporated off under vacuum, (iv) the remaining aqueous phase solution was extracted three times with toluene and the pH of the aqueous solution again adjusted to 7.3-7.4 using 30% sodium hydroxide solution and water was evaporated off under vacuum to obtain a residue, (v) the residue was then
taken up in isopropanol and evaporated under vacuum, (vi) the residue is further taken up in isopropanol and the precipitated mineral salts were filtered off; the filtrate obtained was evaporated under vacuum to obtain crude product, (vii) the crude product was dissolved in methanol and crystallized by adding isopropyl acetate, (viii) the suspension was stirred overnight at 20°C, the solid obtained was filtered off and dried to obtain emtricitabine, having yield 31%. The US'885 Patent utilizes salt of FCME, such as oxalate salt of FCME for the preparation of emtricitabine. Moreover, after the reaction completion, pH of the whole reaction mixture was adjusted to 4 using 37% hydrochloric acid and then the organic solvents were evaporated off under vacuum, this leads to the handling of large volume of reaction mixture. Also the step (viii) of the above discussed process requires overnight stirring for separation of the product, emtricitabine with a yield of only 31%. Therefore, the process of the US'885 Patent, as discussed herein above for the preparation of emtricitabine is disadvantageous for the industrial manufacturing of emtricitabine.
US Patent Application Publication No. 2010/0256372 discloses a process for the preparation of emtricitabine of formula I comprising the steps of: (i) reducing the compound FCME of formula II employing the reducing agent in the presence of organic solvents to obtain reaction mixture containing emtricitabine, (ii) followed by insitu salification of emtricitabine using an organic acid or a mineral acid in the presence of an organic solvent to yield organic or mineral acid salt of emtricitabine, such as hydrochloride, hydrobromide or methane sulfonate salt of emtricitabine, (iii) neutralizing the product obtained in step (ii) using an organic amine bases in the presence of organic solvent to yield emtricitabine. The process disclosed in the US '372 patent appln. involves three steps process for the preparation of emtricitabine, involving first step of reduction of FCME to emtricitabine, second step of isolation of salt of emtricitabine from the reaction mixture and third step of conversion of salt of emtricitabine to emtricitabine free base, thereby making the process lengthy and costly. Also the process involves lengthy work-up process for the isolation of salt of emtricitabine as well as emtricitabine free base from the reaction mixture, thus renders the process for preparation of emtricitabine industrially not viable.
The process for the preparation of emtricitabine of formula I can be improved particularly in terms of industrial applicability, by providing cost-effective and simple isolation process for emtricitabine from the reaction mixture, that would result in obtaining emtricitabine in good yield and having high purity. The process for the isolation of emtricitabine disclosed in the cited prior art references mostly suggests insitu salification of emtricitabine for the isolation of the salt of emtricitabine such as monosalicylate, oxalate, hydrochloride, hydrobromide, methane sulfonate etc. from its reaction mixture and subsequent neutralization of salt of emtricitabine to yield emtricitabine free base, rather than directly isolating emtricitabine free base from the reaction mixture. The prior art process for the preparation emtricitabine necessitates isolation of emtricitabine from the reaction mixture as a salt of emtricitabine for the reason being emtricitabine is highly soluble in water and other polar solvents, thereby making isolation of emtricitabine from the reaction mixture difficult. As a result most of the prior art processes involve isolation of insoluble salt of emtricitabine from the reaction mixture containing water or other polar solvent as a solvent and subsequent conversion of salt of emtricitabine to emtricitabine free base. Thus, there is need to develop a single step process for the preparation of emtricitabine from FCME of formula II, avoiding preparation of any intermediate salt of emtricitabine, which is simple, cost-effective and industrially applicable.
The inventors of the present invention have now found that emtricitabine of formula I can be obtained in good yield and high purity from (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II through an improved process, which although involves use of sodium borohydride as a reducing agent in the presence of phosphate buffer, provides a single step process for the preparation of emtricitabine from FCME and avoids preparation of any salt of emtricitabine as an intermediate step, which in turn reduces the number of steps. Thus, the present invention provides a simple, cost-effective, industrially applicable process for the isolation of emtricitabine, which is used as a nucleoside reverse transcriptase inhibitor.
OBJECTS OF THE INVENTION
An object of the present invention is to provide a process for the isolation of emtricitabine of formula I from a reaction mixture obtained by reaction of the compound of formula II with a reducing agent in the presence of phosphate buffer and a solvent, resulting in obtaining emtricitabine having yield of > 60% and purity of > 99.8%.
Another object of the present invention is to provide an alternate process for the isolation of emtricitabine of formula I from a reaction mixture obtained by reaction of the compound of formula II with a reducing agent in the presence of phosphate buffer and a solvent, resulting in obtaining emtricitabine having purity > 99.8%.
Yet another object of the present invention is to provide a process for the isolation of emtricitabine of formula I, which is simple and cost-effective, and avoids isolation of salts of emtricitabine.
Still another object of the present invention is to provide a process for the isolation of emtricitabine of formula I, which is industrially applicable.
SUMMARY OF THE INVENTION
In accordance with the aspect of the present invention, there is provided a process for the isolation of 4-amino-5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5- yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture; comprising the steps of:
a) obtaining the reaction mixture by reacting (lR,2S,5R)-menthyl-(2S,5R)-5-(4- amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to 20°C,
b) separating the polar solvent from the resulting reaction mixture,
c) adding an aqueous acid to the mixture of step (b) to adjust pH of the reaction mixture to 8.0 to 8.5,
d) distilling out the polar solvent from the mixture of step (c) to obtain an aqueous solution,
e) extracting the aqueous solution of step (d) with a non-polar solvent and distilling out the aqueous solution to obtain a liquid,
f) diluting the liquid of step (e) with a polar solvent and distilling out the solvent to remove the traces of water from the reaction mixture,
g) diluting the mixture of step (f) with a polar solvent and refluxing the reaction mixture at a temperature of 85°C to 90°C for 30 minutes to precipitate out an inorganic residue,
h) filtering off the inorganic residue obtain in step (g) and distilling out the filtrate to obtain a residue,
i) diluting the residue of step (h) with an alcohol and heating it for 30 minutes at a temperature of 85°C to 90°C, and
j) cooling the reaction mixture of step (i) at a temperature of 10°C to 15°C and isolating the precipitated emtricitabine by filtration, having purity of >99.8 .
In accordance with another aspect of the present invention, there is provided an alternate process for the isolation of 4-amino-5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)- l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture; comprising the steps of:
(i) obtaining the reaction mixture by reacting (lR,2S,5R)-menthyl-(2S,5R)-5-(4- amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II with a solution of sodium borohydride in aqueous
sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to 20°C,
(ii) separating the polar solvent from the resulting reaction mixture,
(iii) adding an aqueous acid to the mixture of step (ii) to adjust pH of the reaction mixture to 8.0 to 8.5,
(iv) distilling out the polar solvent from the mixture of step (iii) to obtain an aqueous solution,
(v) extracting the aqueous solution of step (iv) with a non-polar solvent,
(vi) diluting the aqueous solution of step (v) with polar solvent at a temperature ranging from 15°C to 35°C, and
(vii) cooling the reaction mixture of step (vi) slowly at a temperature of 5°C to 10°C over a period of 6 to 12 hours and isolating the precipitated emtricitabine by filtration, having purity of > 99.8%.
In accordance with yet another aspect of the present invention, the process for isolation of emtricitabine of formula I overcomes the disadvantages associated with the processes disclosed in the cited prior art, which mainly concerns with the isolation of salt of emtricitabine from the reaction mixture. The reason being emtricitabine is highly soluble in the solvents used during the reaction and thereby facing the difficulties to isolate pure emtricitabine free base from the reaction mixture. The present invention provides a solution to this problem by providing the processes that involve use of non-hazardous and inexpensive reducing agents such as sodium
borohydride, thereby avoiding use of hazardous reducing agents such as lithium aluminum hydride (LAH) as discussed in the prior art.
In accordance with still another aspect of the present invention, the process for the isolation of emtricitabine of formula I from the reaction mixture provides the desired emtricitabine free base with good yield and high purity, without use of any lengthy isolation process and hazardous reducing agents, thereby making the process of the isolation of emtricitabine simple, cost-effective and industrially applicable.
DESCRIPTION OF THE INVENTION
The present invention relates to a process for the isolation of 4-amino-5-fluoro-l- [(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture comprising the steps of:
Formula I
a) obtaining the reaction mixture by reacting (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II
Formula II
with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar
solvent based on the compound of formula II, at a temperature of 15°C to 20°C,
b) separating the polar solvent from the resulting reaction mixture,
c) adding an aqueous acid to the mixture of step (b) to adjust pH of the reaction mixture to 8.0 to 8.5,
d) distilling out the polar solvent from the mixture of step (c) to obtain an aqueous solution,
e) extracting the aqueous solution of step (d) with a non-polar solvent and distilling out the aqueous solution to obtain a liquid,
f) diluting the liquid of step (e) with a polar solvent and distilling out the solvent to remove the traces of water from the reaction mixture,
g) diluting the mixture of step (f) with a polar solvent and refluxing the reaction mixture at a temperature of 85°C to 90°C for 30 minutes to precipitate out an inorganic residue,
h) filtering off the inorganic residue obtain in step (g) and distilling out the filtrate to obtain a residue,
i) diluting the residue of step (h) with an alcohol and heating it for 30 minutes at a temperature of 85°C to 90°C, and
j) cooling the reaction mixture of step (i) at a temperature of 10°C to 15°C and isolating the precipitated emtricitabine by filtration, having purity of >99.8 .
In an embodiment of the present invention, in the step (a) of the process said FCME of formula II is reacted with sodium borohydride as a reducing agent in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent to obtain the reaction mixture.
In accordance with the embodiment of the present invention, the reaction is carried out in the presence of a polar solvent selected from alcohols, ethers or a mixture thereof. The ether used for the reaction as a solvent is selected from tetrahydrofuran, dioxane, diethyl ether or dimethyl ether or a mixture thereof. The alcohol used for the reaction as a solvent is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof, preferably the solvent used for the reaction is isopropyl alcohol.
In accordance with the embodiment of the present invention, the process involves: taking isopropyl alcohol in a reaction vessel and a solution of dipotassium hydrogen phosphate in water is charged. To this reaction mixture is then charged a solution of FCME of formula II dissolved in 9 volumes of isopropyl alcohol, followed by a solution of sodium borohydride in aqueous sodium hydroxide and stirred at temperature of 15°C to 20°C for 4 to 5 hours to obtain the reaction mixture. The addition of solution of sodium borohydride in aqueous sodium hydroxide to the reaction mixture is carried out with caution, since addition of sodium borohydride to the reaction mixture at a fast rate may result in defluorination of emtricitabine of formula I generating lamivudine as an impurity. Thus, in the present invention, the solution of sodium borohydride in aqueous sodium hydroxide is added slowly to the reaction mixture over a period of 1 to 1.5 hours. It has been indicated herein above that the inventors of the present invention performed numerous experiments to develop an efficient process for the isolation of emtricitabine of formula I from the reaction mixture obtained by reacting FCME of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to 20°C. The process pertaining to step (a) of the present invention was carried out at varying temperature conditions with the constant volume of a polar solvent. The results of effect of varying temperature on the time required for the completion of reaction and the purity of the emtricitabine obtained are presented in the following Table- 1.
Table-1:
Ex. No. = Experiment Number
The results presented in the above Table-1 are discussed herein below:
> As indicated in Ex. No. 1, when the reaction mixture obtained by reacting FCME of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 5°C-10°C, it results in completion of the reaction only after 20 hours and the emtricitabine obtained having purity 95%.
> As indicated in Ex. No. 2, when the reaction mixture obtained by reacting FCME of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C-20°C, it results in completion of reaction within 4 hours with improvement in the purity of emtricitabine of formula I, i.e. 99.8% purity [with respect to Ex. No. 1].
> As indicated in Ex. No. 3, when the reaction mixture obtained by reacting FCME of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 25°C-30°C, it results in completion of reaction within 4 hours with decrease in the purity of emtricitabine of formula I, i.e. 91% purity, as at a temperature of 25°C-30°C the reaction generates 6% lamivudine as an impurity [with respect to Ex. No. 2].
Further, the inventors of the present invention studied the effect of varying volume of polar solvent used in step (a) of the process on the time required for the completion of reaction. In this experiment the temperature condition was maintained constant. The results of the experiments are presented in the following Table-2.
Table-2:
Ex. Solvent Reaction Time in Reaction completion in
No. volume temperature hours %
1. 9 15°C-20°C 4 99.2
2. 5 15°C-20°C 4 70.6
3. 5 15°C-20°C 8 80
4. 5 15°C-20°C 18 92.7
5. 5 15°C-20°C 25 97.8
6. 5 15°C-20°C 35 99.4
Ex. No. = Experiment Number
The results presented in the above Table-2 are discussed herein below:
As indicated in Ex. No. 1, when the reaction mixture obtained by reacting FCME of formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C-20°C, it results in about 99% completion of reaction within 4 hours.
As indicated in Ex. No. 2 to 6, it is observed that when the reaction mixture is obtained by reacting FCME of formula II with solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 5 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C-20°C, the reaction reaches to completion only after 35 hours. Thus, it is evident from the results presented in table-2 that when less than 9 volumes of polar solvent (Ex. 2) the reaction does not lead to completion in 4 hours as was observed when the reaction was carried out using 9 volumes of the polar solvent (Ex. 1). Ultimately it can be observed from the Ex. 3 to Ex. 6 that it requires at least 35 hours for completion of the reaction when 5 volumes of the polar solvent is used.
In another embodiment of the present invention, in step (b) of the process, the aqueous and organic layers formed in the reaction mixture are separated using a separation flask. The organic layer containing the polar solvent such as isopropyl alcohol is used as such for the next step. At this stage the pH of the organic layer is approximately 11.2.
In yet another embodiment of the present invention, in the step (c) of the process, the reaction mixture obtained in step (b) is adjusted to pH of 8.0 to 8.5 using an aqueous acid.
In accordance with yet another embodiment of the present invention, the aqueous acid used in the reaction is a mineral acid selected from hydrochloric acid and hydrobromic acid, or a weak organic acid selected from formic acid, acetic acid or oxalic acid or a mixture thereof, preferably hydrochloric acid is used as an aqueous acid in the reaction. The hydrochloric acid used in the reaction to adjust the pH of the reaction mixture to 8.0 to 8.5 is 20% (v/v) dilute hydrochloric acid.
In yet another embodiment of the present invention, in the step (d) of the process, the reaction mixture obtained in step (c) is distilled out at a temperature of 40°C to 45°C under vacuum to obtain an aqueous solution.
In yet another embodiment of the present invention, in the step (e) of the process, the aqueous solution obtained in step (d) is extracted with a non-polar solvent. In accordance with yet another embodiment of the present invention, the non-polar solvent used for the step (e) of the process is selected from hexane, cyclohexane, toluene, benzene, dioxane, chloroform or diethyl ether or mixture thereof, preferably the non-polar solvent used for the reaction is toluene. In accordance with yet another embodiment of the present invention, the aqueous solution obtained in step (e) is extracted with a non-polar solvent to remove the byproduct menthol from the reaction mixture, which is formed during the reaction.
In accordance with yet another embodiment of the present invention, the aqueous solution obtained after the extraction with non-polar solvent such as toluene, the aqueous solution may optionally be treated with charcoal for 30 minutes at a temperature of 25°C to 30°C and the reaction mixture obtained is filtered off. The filtrate obtained is distilled out under vacuum at 50°C to 60°C to obtain a liquid. In yet another embodiment of the present invention, in the step (f) of the process, the liquid obtained in step (e) is diluted with a polar solvent such as isopropyl alcohol and distill out the polar solvent at a temperature of 50°C to 60°C under vacuum.
In accordance with yet another embodiment of the present invention, the liquid obtained in step (e) is treated with a polar solvent for the complete removal of water content from the reaction mixture. The presence of water in the reaction mixture retains the water soluble inorganic residue in the reaction mixture, which will get extracted along with the product in the subsequent steps. As a result of which the product, emtricitabine of formula I may fail in the sulfated ash test. Moreover, emtricitabine has relatively good solubility in water, the presence of traces water in the reaction mixture may also result in loss of yield of the product. Thus, in order to ensure complete removal of water content from the reaction mixture, the reaction mixture may be treated with a polar solvent for multiple numbers of times. The polar solvent obtained is distilled out from the reaction mixture at a temperature of 50°C to 60°C under vacuum to obtain a thick solid. At this stage the thick solid obtained is analyzed for moisture content and the limit for moisture content is not more than (NMT) 1.5%.
In yet another embodiment of the present invention, in the step (g) of the process, the thick solid obtained in step (f) is diluted with a polar solvent at a temperature of 50°C to 60°C and reflux the reaction mixture at a temperature of 85°C to 90°C for 30 minutes, the product, emtricitabine of formula I gets extracted into the polar solvent and the undissolved inorganic residue precipitated out from the reaction mixture.
In still another embodiment of the present invention, in step (h) of the process, the reaction mixture obtained in step (g) is cooled and filtered to remove undissolved inorganic residue from the reaction mixture. The filtrate obtain is then distilled out at a temperature of 45°C to 50°C under vacuum to obtain a residue.
In accordance with still another embodiment of the present invention, the residue obtained in step (h) may diluted with alcohol and distilled out alcohol at a temperature of 45°C to 50°C under vacuum to obtain the residue. At this stage of the reaction, the solvent of the reaction mixture is changed from isopropyl alcohol to ethanol.
In accordance with still another embodiment of the present invention, the alcohol used in the reaction is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof, preferably said alcohol is ethanol.
In still another embodiment of the present invention, in the step (i) of the process, the residue obtained at stage (h) is diluted with 2.5 volume of alcohol and the reaction mixture is heated at a temperature of 85°C to 90°C for 30 minutes.
In still another embodiment of the present invention, in the step (j) of the process, the reaction mixture obtained in step (i) is cooled to a temperature of 10°C to 15°C and the reaction mixture is stirred at a temperature of 10°C to 15°C for 4 hours to precipitate the product, emtricitabine of formula I. The product obtained under vacuum is filtered and washed with ethanol. The product is dried under vacuum at a temperature of 45°C to 50°C. The emtricitabine obtained by using the above discussed process having yield > 60% and purity > 99.8%.
The present invention also relates to an alternate process for the isolation of 4-amino- 5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)-l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture comprising the steps of:
Formula I
obtaining the reaction mixture by reacting (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II
H3C "CH3
Formula II
with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to 20°C,
(ii) separating the polar solvent from the resulting reaction mixture,
(iii) adding an aqueous acid to the mixture of step (ii) to adjust pH of the reaction mixture to 8.0 to 8.5,
(iv) distilling out the polar solvent from the mixture of step (iii) to obtain an aqueous solution,
(v) extracting the aqueous solution of step (iv) with a non-polar solvent,
(vi) diluting the aqueous solution of step (v) with polar solvent at a temperature ranging from 15°C to 35°C, and
(vii) cooling the reaction mixture of step (vi) slowly at a temperature of 5°C to 10°C over a period of 6 to 12 hours and isolating the precipitated emtricitabine by filtration having purity of > 99.8%.
In an embodiment of the present invention, in the step (i) of the process said FCME of formula II is reacted with sodium borohydride as a reducing agent in the presence of dipotassium hydrogen phosphate and 9 volume of polar solvent to obtain a reaction mixture.
In accordance with the embodiment of the present invention, the reaction is carried out in the presence of a polar solvent selected from alcohols, ethers or a mixture thereof. The ether used for the reaction as a solvent is selected from tetrahydrofuran, dioxane, diethyl ether or dimethyl ether or a mixture thereof. The alcohol used for the reaction as a solvent is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof, preferably the solvent used for the reaction is isopropyl alcohol.
In accordance with the embodiment of the present invention, the process involves: taking isopropyl alcohol in a reaction vessel and a solution of dipotassium hydrogen phosphate in water is charged. To this reaction mixture is then charged a solution of FCME of formula II dissolved in 9 volumes of isopropyl alcohol, followed by solution of sodium borohydride in aqueous sodium hydroxide and stirred at temperature of 15°C to 20°C for 4 to 5 hours to obtain the reaction mixture. The addition of solution of sodium borohydride in aqueous sodium hydroxide to the reaction mixture is carried out with caution, since addition of sodium borohydride to the reaction mixture at a fast rate may result in defluorination of emtricitabine of formula I generating lamivudine as an impurity. Thus, in the present invention, the solution of sodium borohydride in aqueous sodium hydroxide is added slowly to the reaction mixture over a period of 1 to 1.5 hours. In another embodiment of the present invention, in step (ii) of the process, the aqueous and organic layers formed in the reaction mixture are separated using a separation flask. The organic layer containing the polar solvent such as isopropyl alcohol is used as such for the next step. At this stage the pH of the organic layer is approximately 11.2.
In yet another embodiment of the present invention, in the step (iii) of the process, the reaction mixture obtained in step (ii) is adjusted to pH of 8.0 to 8.5 using an aqueous acid. In accordance with yet another embodiment of the present invention, the aqueous acid used in the reaction is a mineral acid selected from hydrochloric acid and hydrobromic acid, or a weak organic acid selected from formic acid, acetic acid or oxalic acid or a mixture thereof, preferably hydrochloric acid is used as an aqueous acid in the reaction. The hydrochloric acid used in the reaction to adjust the pH of the reaction mixture to 8.0 to 8.5 is 20% (v/v) dilute hydrochloric acid.
In yet another embodiment of the present invention, in the step (iv) of the process, the reaction mixture obtained in step (iii) is distilled out at a temperature of 40°C to 45°C under vacuum to obtain an aqueous solution. In yet another embodiment of the present invention, in the step (v) of the process, the aqueous solution obtained in step (iv) is extracted with a non-polar solvent.
In accordance with yet another embodiment of the present invention, the non-polar solvent used for the step (v) is selected from hexane, cyclohexane, toluene, benzene, dioxane, chloroform or diethyl ether or mixture thereof, preferably the non-polar solvent used for the reaction is toluene.
In accordance with yet another embodiment of the present invention, the aqueous solution obtained in step (iv) is extracted with a non-polar solvent to remove the by- product menthol from the reaction mixture, which is formed during the reaction.
In yet another embodiment of the present invention, in the step (vi) of the process, the aqueous solution obtained in step (v) may optionally stirred at a temperature of 15°C to 35°C for 15 to 20 minutes. The aqueous solution is then diluted with 2 volumes of polar solvent, such as isopropyl alcohol at a temperature of 15°C to 35°C.
In still another embodiment of the present invention, in the step (vii) of the process, the reaction mixture slowly cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours to precipitate the product, emtricitabine of formula I. The precipitated product is then filtered and washed with isopropyl alcohol. The product dried under vacuum at a temperature of 45°C to 50°C, having yield > 50% and purity > 99.8%.
It has been indicated herein above that the inventors of the present invention performed numerous experiments to develop an efficient process for the isolation of emtricitabine of formula I. The process discussed herein above pertaining to step (vi) and step (vii) of the present invention was carried out under different reaction conditions particularly, the results of effect of varying volumes of solvent at different
temperature conditions and at varying time duration on the yield of emtricitabine are presented in the following Table-3.
Table-3:
Ex. No. = Experiment Number
The results presented in the above Table-3 are discussed herein below:
> As discussed in Ex. No. 1, when the aqueous solution obtained in step (v) is diluted with 1 volume of a polar solvent, based on the compound of formula II, at a temperature of 15°C to 35°C and then the reaction mixture is slowly cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours, it results with 30% yield of emtricitabine of formula I.
> As discussed in Ex. No. 2, when the aqueous solution obtained in step (v) is diluted with 2 volume of polar solvent, based on the compound of formula II, at a temperature of 15°C to 35°C and then the reaction mixture is slowly cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours, it results with increase in the yield of emtricitabine of formula I, i.e. emtricitabine is obtained in a yield of 50% (with respect to Ex. No. 1).
> As discussed in Ex. No. 3, when the aqueous solution obtained in step (v) is diluted with 2 volume of polar solvent, based on the compound of formula II, at a temperature of 15°C to 35°C and then the reaction mixture is filtered at a temperature of ~30°C, avoiding cooling of the reaction mixture to a temperature of 5°C-10°C over a period of 6 to 12 hours, it results with significant decrease in the yield of emtricitabine of formula I i.e. emtricitabine is obtained in a yield of only 25% (with respect to Ex. No. 2)
> As discussed in Ex. No. 4, when the aqueous solution obtained in step (v) is diluted with 10 volumes of polar solvent, based on the compound of formula II, at a temperature of 15°C to 35°C and then the reaction mixture slowly
cooled to a temperature of 5°C to 10°C over a period of 6 to 12 hours, it results with decrease in the yield of emtricitabine of formula I i.e. emtricitabine is obtained in a yield of 32% (with respect to Ex. No. 2) In accordance with the present invention the emtricitabine of formula I is isolated from the reaction mixture according to the process described herein below:
To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, charged dipotassium hydrogen phosphate and water and stirred the reaction mixture for 15 minutes. Then charged polar solvent, such as isopropyl alcohol to the reaction vessel and further stirred the reaction mixture for 15 minutes and the reaction mixture obtained is cooled to a temperature of 15°C to 20°C. To the reaction mixture then added FCME of formula II dissolved in isopropyl alcohol and stirred the reaction mixture at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride dissolved in aqueous sodium hydroxide added dropwise to the reaction mixture over the period of l-1.5hours. Stir the reaction mixture for 4 - 5hours and the two layers formed were separated. To the separated organic layer 20% (v/v) dilute hydrochloric acid is added dropwise to adjust the pH of the reaction mixture to 8.0-8.5 and stirred the reaction mixture for 15 minutes. The reaction mixture is then distilled out to obtain an aqueous solution. The aqueous solution obtained is extracted with non-polar solvent such as toluene and treated with charcoal and filtered. The filtrate is then distilled out to obtain a liquid, diluted the liquid with polar solvent and distilled out the polar solvent at a temperature of 50°C to 60°C. Dilute the reaction mixture with 2nd lot of polar solvent and distilled out. Further diluted the reaction mixture with 3 rd lot of polar solvent and reflux the reaction mixture to a temperature of 85°C to 90°C for 30 minutes to obtain the residue. Cool the reaction mixture and filter, the filtrate obtained is then distilled out at a temperature of 45°C to 50°C to obtain the residue. The residue obtained is then diluted with alcohol such as ethanol and distilled out the alcohol at a temperature of 45°C to 50°C. The reaction mixture further diluted with 2nd lot of alcohol and the reaction mixture obtained is heated at a temperature of 85°C to 90°C for 30 minutes and cool the reaction mixture at a temperature of 10°C to 15°C to obtain the product, emtricitabine. Filter the product obtained and wash with ethanol. Dry the product under vacuum. The emtricitabine obtained from the above discussed process having yield > 60% and purity > 99.8%.
In accordance with the present invention the emtricitabine of formula I is isolated from the reaction mixture according to the alternate process described herein below: To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, charged dipotassium hydrogen phosphate and water and stirred the reaction mixture for 15 minutes. Then charged polar solvent, such as isopropyl alcohol to the reaction vessel and further stirred the reaction mixture for 15 minutes and the reaction mixture obtained is cooled to a temperature of 15°C to 20°C. To the reaction mixture then added FCME of formula II dissolved in isopropyl alcohol and stirred the reaction mixture at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride dissolved in aqueous sodium hydroxide added dropwise to the reaction mixture over the period of l-1.5hours. Stir the reaction mixture for 4 - 5hours and the two layers formed were separated. To the separated organic layer 20% (v/v) dilute hydrochloric acid is added dropwise to adjust the pH of the reaction mixture to 8.0-8.5 and stirred the reaction mixture for 15 minutes. The reaction mixture is then distilled out to obtain an aqueous solution. The aqueous solution obtained is extracted with non-polar solvent such as toluene The aqueous layer obtained is diluted with a polar solvent at a temperature of 15°C to 20°C and slowly cooled to a temperature of 5°C to 10°C over a period of 6-12 hours to obtain the product, emtricitabine. Filter the product obtained and wash with polar solvent. The emtricitabine obtained from the above discussed process having yield > 50% and purity > 99.8%.
It is thus possible by the way of the present invention to achieve the much desired process for the isolation of emtricitabine of formula I with good yield and high purity using a simple, cost-effective and industrially applicable process.
The starting material of the process, (lR,2S,5R)-menthyl-(2S,5R)-5-(4-amino-5- fluoro-2-oxo-l(2H)-pyrimidinyl)-l,3-oxathiolane-2-carboxylate (FCME) of formula II is a known compound and can be prepared by a person skilled in the art by following the process disclosed in the literature. For example, the compound of formula II may be prepared by following the process disclosed in the US Patent No. 5,696,254, which is incorporated herein by reference. The process involves reaction of 5-fluorocytosine with 2,,4,6-collidine and t-butyldimethylsilyl trifluoromethane sulfonate in the presence of dichloromethane, the resultant reaction mixture was
stirred for 15 minutes to obtain the clear solution. A separately prepared solution of (lR,2S,5R)-menthyl-5R-acetoxy-l,3-oxathiolane-2S-carboxylate in dichloromethane was added to the clear solution, followed by iodotrimethylsilane and stirred for 3 hours. Further reaction work-up yields the product FCME of formula II.
The following examples which fully illustrate the practice of the preferred embodiment of the present invention are intended to be illustrative purpose only and should not be constructed in any way to limit the scope of the present invention. EXAMPLES
Example 1:
To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, dipotassium hydrogen phosphate (147. lg) and water (330ml) was charged and stirred the reaction mixture for 15 minutes. To the reaction mixture then charged isopropyl alcohol (800ml) and further stirred for 15 minutes and cooled the reaction mixture to a temperature of 15°C to 20°C. Then charged FCME (lOOg) dissolved in isopropyl alcohol (100ml) to the reaction mixture and stirred at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride (26g) dissolved in aqueous sodium hydroxide was added dropwise to the reaction mixture over the period of 1-1.5 hours. The reaction mixture further stirred for 4 - 5 hours and the two layers formed were separated. To the separated organic layer 20% (v/v) dilute hydrochloric acid (4.6ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was further stirred for 15 minutes. The reaction mixture was then distilled out and the aqueous solution obtained was extracted with toluene (200ml). The reaction mixture then treated with charcoal (5g) and filtered. The filtrate obtained was distilled out to obtain a liquid and diluted with isopropyl alcohol (200ml), distilled out the isopropyl alcohol. Diluted the reaction mixture again with isopropyl alcohol (200ml) and distilled out. The reaction mixture further diluted with isopropyl alcohol (400ml) and refluxed the reaction mixture to a temperature of 85°C to 90°C for 30 minutes to obtain the residue. Cooled the reaction mixture and filtered, the filtrate obtained was then distilled out to obtain the residue. Diluted the residue obtained with ethanol (100ml) and distilled out. The reaction mixture again diluted with ethanol (200ml) and heated at a temperature of 85°C to 90°C for 30 minutes and
then cooled at a temperature of 10°C to 15°C to precipitate the product, emtricitabine. The resulting product was filtered and washed with ethanol. Dry the product under vacuum.
Yield = 62%
Assay = 98.7%
Purity = 99.86%
Example 2: To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, dipotassium hydrogen phosphate (73.8g) and water (165ml) was charged and stirred the reaction mixture for 15 minutes. To the reaction mixture then charged isopropyl alcohol (350ml) and further stirred for 15 minutes and cooled the reaction mixture to a temperature of 15°C to 20°C. Then charged FCME (50g) dissolved in isopropyl alcohol (100ml) to the reaction mixture and stirred at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride (13g) dissolved in aqueous sodium hydroxide was added dropwise to the reaction mixture over the period of 1-1.5 hours. The reaction mixture further stirred for 4 - 5 hr and the two layers formed were separated. To the separated organic layer 20% (v/v) dilute hydrochloric acid (1.8ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was further stirred for 15 minutes. The reaction mixture was distilled out and the aqueous solution obtained was extracted with toluene (100ml). The reaction mixture then treated with charcoal (2.5g) and filtered. The filtrate obtained was distilled out to obtain a liquid and diluted with isopropyl alcohol (50ml), distilled out the isopropyl alcohol. The reaction mixture was again diluted with isopropyl alcohol (100ml) and distilled out. The reaction mixture was further diluted with isopropyl alcohol (250ml) and refluxed the reaction mixture to a temperature of 85°C to 90°C for 30 minutes to obtain the residue. Cooled the reaction mixture and filtered, the filtrate obtained was then distilled out to obtain the residue. The residue obtained was diluted with ethanol (50ml) and distilled out. The reaction mixture again diluted with ethanol (150ml) and heated at a temperature of 85°C to 90°C for 30 minutes and then cooled at a temperature of 10°C to 15°C to precipitate the product, emtricitabine. The resulting product was filtered and washed with ethanol. Dry the product under vacuum.
Yield = 60%
Assay = 99.9%
Purity = 99.91% Example 3:
To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, dipotassium hydrogen phosphate (147. lg) and water (330ml) was charged and stirred the reaction mixture for 15 minutes. To the reaction mixture then charged isopropyl alcohol (900ml) and further stirred for 15 minutes and cooled the reaction mixture to a temperature of 15°C to 20°C. Then charged FCME (lOOg) dissolved in isopropyl alcohol to the reaction mixture and stirred at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride (23.5g) dissolved in aqueous sodium hydroxide was added dropwise to the reaction mixture over the period of 1-1.5 hours. The reaction mixture further stirred for 4 - 5 hours and the two layers formed were separated. To the separated organic layer 20% (v/v) dilute hydrochloric acid (4ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was stirred for 15 minutes. The reaction mixture was distilled out and the aqueous solution obtained was extracted with toluene (200ml). The aqueous solution obtained was then diluted with isopropyl alcohol (200ml) at a temperature of 15°C to 20°C and slowly cooled to a temperature of 5°C to 10°C over a period of 6- 12 hours to obtain the product, emtricitabine. The product was filtered and washed with isopropyl alcohol.
Yield = 50%
Purity = 99.91%
Example 4:
To a 2.0 L capacity jacketed flask equipped with a mechanical stirrer, thermometer pocket, dipotassium hydrogen phosphate (73.6g) and water (165ml) was charged and stirred the reaction mixture for 15 minutes. To the reaction mixture then charged isopropyl alcohol (350ml) and further stirred for 15 minutes and cooled the reaction mixture to a temperature of 15°C to 20°C. Then charged FCME (50g) dissolved in isopropyl alcohol to the reaction mixture and stirred at a temperature of 15°C to 20°C for 1 hour. A separately prepared solution of sodium borohydride (13g) dissolved in
aqueous sodium hydroxide was added dropwise to the reaction mixture over the period of 1-1.5 hours. The reaction mixture further stirred for 4 - 5 hours and the two layers formed were separated. To the separated organic layer 20% (v/v) dilute hydrochloric acid (2ml) was added dropwise to adjust the pH to 8.0-8.5 and the reaction mixture was stirred for 15 minutes. The reaction mixture was distilled out and the aqueous solution obtained was extracted with toluene (100ml). The aqueous solution obtained was then diluted with isopropyl alcohol (100ml) at a temperature of 15°C to 20°C and slowly cooled to a temperature of 5°C to 10°C over a period of 6- 12 hours to obtain the product, emtricitabine. The product was filtered and washed with isopropyl alcohol.
Yield = 53%
Purity = 99.69%
Details For Analytical Method Of Analysis:
HPLC column Hypersil BDS, 4.6mm x 10cm, 5μιη packing LI
Detector UV 280nm
Mobile phase Acetonitrile and buffer (1:9)
Buffer preparation 2.72g/L of monobasic potassium phosphate and 4.32g/L of 1- octanesulfonic acid sodium salt in water. Adjust with 20% phosphoric acid to a pH of 2.2.
Standard solution 0.1 mg/ml of USP emtricitabine RS in mobile phase.
Sample solution 0.1 mg/ml of mobile phase
Injection volume ΙΟμί
Column temperature 30°C
Flow rate 1.5ml/minutes
Run time 2.5 times the retention time of emtricitabine
Claims
CLAIM:
>rocess for the isolation of 4-amino-5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)- oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a :tion mixture comprising the steps of:
Formula I
a) obtaining the reaction mixture by reacting (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II
with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to b) separating the polar solvent from the resulting reaction mixture,
c) adding an aqueous acid to the mixture of step (b) to adjust pH of the reaction mixture to 8.0 to 8.5,
d) distilling out the polar solvent from the mixture of step (c) to obtain an aqueous solution,
extracting the aqueous solution of step (d) with a non-polar solvent and distilling out the aqueous solution to obtain a liquid,
diluting the liquid of step (e) with a polar solvent and distilling out the solvent to remove the traces of water from the reaction mixture,
diluting the mixture of step (f) with a polar solvent and refluxing the reaction mixture at a temperature of 85°C to 90°C for 30 minutes to precipitate out an inorganic residue,
filtering off the inorganic residue obtain in step (g) and distilling out the filtrate to obtain a residue,
diluting the residue of step (h) with an alcohol and heating it for 30 minutes at a temperature of 85°C to 90°C, and
cooling the reaction mixture of step (i) at a temperature of 10°C to 15°C and isolating the precipitated emtricitabine by filtration, having purity of 99.8%.
The process as claimed in claim 1, wherein said polar solvent is selected from alcohols, ethers or a mixtures thereof.
3. The process as claimed in claim 2, wherein said ether is selected from tetrahydrofuran, dioxane, diethyl ether or dimethyl ether or a mixtures thereof.
4. The process as claimed in claim 2, wherein said alcohol is selected from methanol, ethanol or isopropyl alcohol or a mixtures thereof.
5. The process as claimed in claim 4, wherein said polar solvent is isopropyl alcohol.
6. The process as claimed in claim 1, wherein said aqueous acid is a mineral acid selected from hydrochloric acid or hydrobromic acid, or a weak organic acid selected from formic acid, acetic acid or oxalic acid or a mixture thereof. 7. The process as claimed in claim 6, wherein said aqueous acid is hydrochloric acid.
The process as claimed in claim 7, wherein said hydrochloric acid is 20% (v/v) dilute hydrochloric acid.
The process as claimed in claim 1, wherein said non-polar solvent used in step (e) is selected from hexane, cyclohexane, toluene, benzene, dioxane, chloroform or diethyl ether or a mixture thereof.
10. The process as claimed in claim 9, wherein said non-polar solvent is toluene. 11. The process as claimed in claim 1, wherein in the step (i) said alcohol is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof.
12. The process as claimed in claim 11, wherein said alcohol is ethanol. 13. A process for the isolation of 4-amino-5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)- l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I from a reaction mixture comprising the steps of:
Formula I obtaining the reaction mixture by reacting (lR,2S,5R)-menthyl-(2S,5R)-5- (4-amino-5-fluoro-2-oxo- 1 (2H)-pyrimidinyl)- 1 ,3-oxathiolane-2-carboxylate (FCME) of formula II
H3C "CH3
Formula II with a solution of sodium borohydride in aqueous sodium hydroxide in the presence of dipotassium hydrogen phosphate and 9 volumes of a polar solvent based on the compound of formula II, at a temperature of 15°C to 20°C,
(ii) separating the polar solvent from the resulting reaction mixture,
(iii) adding an aqueous acid to the mixture of step (ii) to adjust pH of the reaction mixture to 8.0 to 8.5,
(iv) distilling out the polar solvent from the mixture of step (iii) to obtain an aqueous solution,
(v) extracting the aqueous solution of step (iv) with a non-polar solvent,
(vi) diluting the aqueous solution of step (v) with polar solvent at a temperature ranging from 15°C to 35°C, and
(vii) cooling the reaction mixture of step (vi) slowly at a temperature of 5°C to 10°C over a period of 6 to 12 hours and isolating the precipitated emtricitabine by filtration having purity of 99.8%.
14. The process as claimed in claim 13, wherein said polar solvent is selected from alcohols, ethers or a mixtures thereof.
15. The process as claimed in claim 14, wherein said ether is selected from tetrahydrofuran, dioxane, diethyl ether or dimethyl ether or a mixture thereof.
16. The process as claimed in claim 14, wherein said alcohol is selected from methanol, ethanol or isopropyl alcohol or a mixture thereof.
17. The process as claimed in claim 16, wherein said polar solvent is isopropyl alcohol.
18. The process as claimed in claim 13, wherein said aqueous acid is a mineral acid selected from hydrochloric acid or hydrobromic acid, or a weak organic acid selected from formic acid, acetic acid or oxalic acid or a mixture thereof.
19. The process as claimed in claim 18, wherein said aqueous acid is hydrochloric acid.
20. The process as claimed in claim 19, wherein said hydrochloric acid is 20% (v/v) dilute hydrochloric acid.
21. The process as claimed in claim 13, wherein said non-polar solvent used in step (v) is selected from hexane, cyclohexane, toluene, benzene, dioxane, chloroform or diethyl ether or a mixture thereof.
22. The process as claimed in claim 21, wherein said non-polar solvent is toluene. 23. A process for the isolation of 4-amino-5-fluoro-l-[(2R,5S)-2-(hydroxymethyl)- l,3-oxathiolan-5-yl]-2(lH)-pyrimidinone (emtricitabine) of formula I as herein described with reference to example 1 to 4.
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WO2014174532A3 (en) * | 2013-04-26 | 2015-02-26 | Laurus Labs Private Limited | An improved process for the preparation of emtricitabine |
CN108409724A (en) * | 2018-06-04 | 2018-08-17 | 安徽帆香料有限公司 | A kind of emtricitabine separation method |
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WO2004085432A1 (en) * | 2003-03-24 | 2004-10-07 | Clariant Life Science Molecules (Italia) Spa | Process and intermediates for preparing emtricitabine |
CN101407513A (en) * | 2008-11-14 | 2009-04-15 | 江苏科本医药化学有限公司 | Method for synthesizing nucleoside analogue |
WO2009084033A2 (en) * | 2007-12-07 | 2009-07-09 | Matrix Laboratories Limited | Process for producing 5-fluoro-1-(2r,5s)-[2-(hydroxymethyl)-1,3-oxathiolan-5-yi]cytosine |
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WO2004085432A1 (en) * | 2003-03-24 | 2004-10-07 | Clariant Life Science Molecules (Italia) Spa | Process and intermediates for preparing emtricitabine |
WO2009084033A2 (en) * | 2007-12-07 | 2009-07-09 | Matrix Laboratories Limited | Process for producing 5-fluoro-1-(2r,5s)-[2-(hydroxymethyl)-1,3-oxathiolan-5-yi]cytosine |
CN101407513A (en) * | 2008-11-14 | 2009-04-15 | 江苏科本医药化学有限公司 | Method for synthesizing nucleoside analogue |
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WO2014174532A3 (en) * | 2013-04-26 | 2015-02-26 | Laurus Labs Private Limited | An improved process for the preparation of emtricitabine |
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