Patent Description:
Influenza is a severe viral infection of the respiratory system, which is responsible for significant morbidity and mortality due to both annual epidemics and predictable pandemics. In the United States alone, <NUM> hospitalisations and <NUM> deaths are recorded per year. In addition, annually the virus affects around <NUM> % of the world population, resulting in about <NUM> deaths.

Oseltamivir phosphate, a compound in the class of compounds referred to as neuraminidase inhibitors (NAls), is used in the treatment and prophylaxis of influenza. It is effective against influenza caused by both the influenza A and influenza B viruses.

There are numerous processes and synthetic routes described in the prior art for the preparation of Oseltamivir phosphate.

However, existing synthesis methodologies for the production of these compounds have essentially been based on standard stirred batch reactor type processes, wherein significant volumes of organic solvents are used. In addition, most of the known processes either employ azide chemistry or protecting group chemistry, both of which introduce inherent limitations, in particular in batch processes.

<CIT> describes such a batch method for a process of producing Oseltamivir phosphate from shikimic acid. The methodology employed is that of a typical batch process, with no suggestion that it may be desirable to attempt to develop a flow synthesis process based on the methods, reagents, and reaction conditions disclosed therein. Further, there is no indication of how the steps disclosed in this application may be adapted and improved to arrive at a flow synthesis method that is superior in both reaction times and reaction yields.

Azide chemistry poses many safety concerns because of its hazardous and highly exothermic nature, which becomes even more pronounced at an industrial scale. Due to these inherent dangers, the process chemist is limited in the reaction parameters that can be employed to maximise reaction efficiency and reaction yield. Protecting group chemistry, on the other hand, generally increases reaction time whilst reducing overall yield, thereby increasing final product cost.

Micro reactor technology (MRT), more recently branded 'flow chemistry', is an emerging technique that enables those working in research and development to rapidly screen reactions utilising continuous flow, leading to the identification of reaction conditions that are suitable for use at a production level. Furthermore, in addition to using conventional reaction methodology, the inherent safety associated with the use of small reactor volumes enables users to employ reaction conditions previously thought to be too hazardous for use within a production environment; such as extreme reaction conditions or the use/generation of 'hazardous' compounds. Consequently, the type of reactions available to the chemist increases through the use of this technology.

Furthermore, in the case of Oseltamivir phosphate, continuous flow synthesis may potentially provide a technology that is efficient enough to enable rapid local manufacture in the event of a pandemic, in particular in developing countries.

The present invention seeks to address some of the shortcomings of the prior art by providing new flow chemistry processes for the production of Oseltamivir.

<CIT>) describes a process for the conversion of shikimic acid to oseltamivir, via the intermediate phosphoramide.

<CIT>) describes an enantioselective process for synthesis of oseltamivir from cis-<NUM>,<NUM>-butene via Sharpless asymmetric epoxidation and diastereoselective Barbier allylation and construction of cyclohexene carboxylic acid ester core through a ring closing metathesis reaction.

According to a first aspect to the present invention there is provided a flow synthesis process for producing a compound of the Formula <NUM> and its pharmaceutically acceptable salts,
<CHM>
the process comprising the steps of:.

Preferably, the process further comprises reacting the compound of Formula <NUM> with phosphoric acid to produce the compound of Formula <NUM>
<CHM>.

Preferably, in step (a) shikimic acid is reacted with (COCl)<NUM> or Amberlyst <NUM>.

In one embodiment, in step (a) shikimic acid is reacted with COCl<NUM> at a temperature of between about <NUM> and about <NUM>.

In another embodiment, in step (a) shikimic acid is reacted with Amberlyst <NUM> at a temperature of between about <NUM> and about <NUM>.

In one embodiment, the reaction in step (b) proceeds at room temperature.

In a preferred embodiment, the reaction in step (b) proceeds under sonication. Preferably, in step (b) the base is present at a concentration of about <NUM> to about <NUM> molar equivalents relative to ethyl shikimate.

In one embodiment, in step (c) the azidation reaction is performed with NaN<NUM> at a concentration of about <NUM> molar equivalents relative to the O-trimesylate of Formula <NUM>.

In a preferred embodiment, in step (c) the azidation reaction is performed with NaN<NUM> at a temperature of about <NUM>.

Preferably, in step (d) the trialkyl phosphite is selected from triethyl phosphite and trimethyl phosphite.

In a preferred embodiment, in step (d) the reaction is performed in acetonitrile at a temperature of between about <NUM> to about <NUM>.

In a preferred embodiment, in step (e) the reaction is performed at a temperature of between about <NUM> to about <NUM>.

Preferably, wherein in step (f) the reaction of the <NUM>-pentyl ether with H<NUM>SO<NUM> is performed at a temperature of between about <NUM> to about <NUM>.

Preferably, in step (g) the reaction is performed at a temperature of between about <NUM> to about <NUM>.

In one embodiment, in step (h) the reaction is performed at about room temperature.

In a preferred embodiment, in step (h) the reaction is performed under sonication.

The invention will now be described in more detail with reference to the following non-limiting embodiments and figures in which:.

The present invention will now be described more fully hereinafter with reference to the accompanying figures, in which some of the non-limiting embodiments of the invention are shown.

The invention as described hereinafter should not be construed to be limited to the specific embodiments disclosed.

Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.

As used herein, throughout this specification and in the claims which follow, the singular forms "a", "an" and "the" include the plural form, unless the context clearly indicates otherwise.

The terminology and phraseology used herein is for the purpose of description and should not be regarded as limiting. The use of the terms "comprising", "containing", "having", "including", and variations thereof used herein, are meant to encompass the items listed thereafter, as well as additional items The present invention provides for a process for producing Oseltamivir and pharmaceutically acceptable salts thereof, in particular to a flow synthesis process for producing this compound. In a particularly preferred embodiment, the invention provides for a flow synthesis process for producing a pharmaceutically acceptable salt of Oseltamivir, (-)-Oseltamivir phosphate commonly referred to under the brand name Tamiflu®.

The invention provides for a continuous flow synthesis process for producing Oseltamivir and pharmaceutically acceptable salts thereof, that does not include the use of rate and yield limiting protecting group chemistry. As a consequence of avoiding protecting group chemistry, we have developed a continuous flow procedure that is high yielding and extremely efficient. In a preferred embodiment of the invention, a <NUM> steps continuous flow procedure towards Oseltamivir phosphate <NUM> is provided having an overall yield of <NUM> % in a total residence time of <NUM>.

Chemicals were supplied by Sigma Aldrich, Merck and Industrial Analytical and were used as received. Anhydrous solvents were supplied by Sigma Aldrich, and maintained by drying over appropriately activated molecular sieves during use.

Column chromatography was performed using Fluka Chemie silica gel <NUM> as the stationary phase, and mixtures of ethyl acetate and hexane of varying polarity were used as the mobile phase. Unless otherwise stated, thin layer chromatography (TLC) was done using Merck Kieselgel <NUM> HF254 aluminium backed TLC plates with mixtures of ethyl acetate and hexane of varying polarity as eluent. TLC visualisation was done by fluorescence on exposure to short wave ultra violet (UV) light (λ <NUM>) in a Camag UV cabinet.

Nuclear magnetic resonance (NMR) spectra were recorded at room temperature as solutions in deuterated chloroform (CDCl<NUM>) or deuterated dimethyl sulfoxide (DMSO-d<NUM>). A Bruker Avance-<NUM> spectrometer (<NUM>) was used to record the spectra and the chemical shifts are reported in parts per million (ppm) with coupling constants in Hertz (Hz). Infra-red spectra were recorded from <NUM> to <NUM>-<NUM> using a Bruker spectrometer and peaks (Vmax) reported in wavenumbers (cm-<NUM>). Melting points of all compounds were determined using a Staurtd Melting Point Apparatus SMP30. High performance liquid chromatography (HPLC) data was obtained using Agilent <NUM> with a UV/Vis detector and Agilent Zorbax C18-column.

HPLC analysis was performed on Agilent Zorbax C18-column (<NUM> x <NUM>, i. <NUM>) ambient temperature using an isocratic system. The mobile phase consisted of <NUM> % acetonitrile and <NUM> % water. The sample injection volume was <NUM>µl, eluted at a flow rate of <NUM>/min and detected at <NUM> with a run time of <NUM>.

The continuous flow systems used in the present invention are described below.

This is a manually operated, 'plug and play' continuous flow reactor system for rapid reaction screening and process optimisation within a micro reactor. The system has a modular set up which facilitates the exchange of components to increase chemical compatibility, number of feed lines or reactor type or volume. It can be used to perform reactions at temperatures ranging from -<NUM> to <NUM> and maximum operating pressure of <NUM> bar (<NUM>×<NUM><NUM> kPa) using very little reagents. This system consists mainly of a Labtrix start unit, thermo-controller, syringe pumps, syringes, tubing and fittings. The start unit holds of the micro reactor. It can be heated or cooled to temperatures between -<NUM> and +<NUM> which is controlled by a thermo-controller. Syringe pump fitted with glass gas tight Leur lock syringes dose reagents into the microreactors. There are twelve different inter-changeable glass micro reactor types available this system which differ in volume and design. These glass reactors have three distinct categories depending on their design and mixing patterns. T-mixer reactors, SOR-mixer reactors and catalyst reactors are the three categories.

The system consisted of a <NUM> i. x <NUM> Omnifit glass column with enhanced PEEK adjustable end fittings. PTFE tubing (<NUM> ID) was used to connect the column reactor to the HPLC pump and from the reactor to the collecting vessel. A peristaltic HPLC pump, Series III (<NUM> pump head) with flow rate range of <NUM> - <NUM>/min was used to dose the liquid reagent through the packed column reactor which was fitted with a <NUM> bar (<NUM><NUM> kPa) back pressure regulator. Uniqsis heating holder was used for heating the glass column reactor.

The system consisted of a Chemyx syringe pump fitted with two <NUM> SGE glass syringes filled with reagents. The two streams of reagents were pumped into a T-mixer (Omnifit labware, Pore size: <NUM> ID, <NUM> - <NUM> OD) which was connected to a <NUM> PTFE coil reactor (<NUM> ID, <NUM> tube length) with a product collection vial downstream. The T-mixer and the PTFE coil reactor were placed into a temperature controlled sonicated water bath. EINS SCI professional ultrasonic bath (<NUM>) was used for sonication.

Shikimic acid <NUM> esterification is the first step in the synthesis of (-)-Oseltamivir phosphate (Scheme <NUM>). Various esterification conditions were investigated to optimise the esterification reaction in Chemtrix Labtrix and Uniqsis packed bed column flow systems. All the solution phase and solid phase esterification were done in a Chemitrix Labtrix system, and Uniqsis packed bed column reactor, respectively.

A Chemtrix Labtrix system was used to perform all solution phase esterification investigations. The system, fitted with a <NUM>µl glass reactor, was used for shikimic acid esterification optimisation in the presence of catalyst. Thionyl chloride, oxalyl chloride, thionyl chloride/DMF, oxalyl chloride/DMF, benzene sulphonic acid (BSA) and p-Toluene sulphonic acid (PTSA) were the various catalyst investigated for shikimic acid esterification. Two syringe pumps were used to pump reagents from two <NUM> SGE Luer lock gas tight glass syringes into the thermally controlled microreactor system which was fitted with a <NUM> bar (<NUM><NUM> kPa) back pressure regulator. Shikimic acid (<NUM>) and catalyst were both dissolved in ethanol and pumped into the flow system separately. Samples were collected and analysed using HPLC method A.

A Uniqsis column reactor packed with a solid catalyst was used all for solid phase esterification investigations (Scheme <NUM>).

A <NUM> ID x <NUM> Uniqsis glass column was packed with Amberlyst <NUM> or Amberlyst (<NUM> bed height, <NUM> reactor volume). The column reactor was heat controlled using Uniqsis heating mantle, and the system was pressurised using a <NUM> bar (<NUM><NUM> kPa) back pressure regulator. A peristaltic HPLC pump was used to pump a solution of shikimic acid (<NUM>) in ethanol into the heated packed bed. Samples were collected and analysed using HPLC method A.

The results and discussions of the detailed investigations on the use of SOCI<NUM>, COCl<NUM>, BSA, PTSA, Amberlyst <NUM> and Amberlyst <NUM> in continuous flow shikimic acid esterification optimisation is reported herein.

Shikimic acid <NUM> esterification with ethanol in the presence of SOCl<NUM> was done in a <NUM>µl glass micro reactor system. Shikimic acid <NUM> (<NUM> in ethanol was treated with SOCl<NUM> (<NUM>, effectively <NUM> equiv. ) at <NUM> and <NUM> residence time. Ethyl shikimate <NUM> was successfully formed (<NUM> %). Doubling residence time resulted in a <NUM> % conversion increase. Increasing temperature to <NUM> and keeping residence time at <NUM> gave <NUM> % conversion. A comprehensive reaction optimisation study was carried out after successful preliminary experiments. The effect of SOCl<NUM> equivalence, reaction temperature and residence time were investigated. An investigating into the effect of SOCl<NUM> molar equivalents on the reaction at <NUM> and <NUM> residence time is shown in <FIG>.

<FIG> illustrates that shikimic acid <NUM> conversion towards ethyl shikimate <NUM> generally increased with increase in SOCl<NUM> concentration. Constant % conversion was achieved at SOCl<NUM> molar equivalents of <NUM> and above. Without thereby wishing to be bound by any particular theory, it is believed that the excess SOCl<NUM> may be necessary because it acts as a catalyst and a water scavenger. At this determined optimum SOCl<NUM> molar equivalence, the effect of residence time and temperature was investigated (<FIG>).

Generally, shikimic acid <NUM> conversion towards ethyl shikimate <NUM> increased with increase in residence time and temperature (<FIG>). However, at residence time above <NUM>, further increase in residence time had limited effect on shikimic acid conversion. Furthermore, at temperatures above <NUM>, shikimic acid conversion decreased. Pressurising the continuous flow system allowed us to investigate reaction temperatures above the boiling point of the solvent/reactant ethanol.

The optimum conditions for this esterification were found to be equimolar equivalents of shikimic acid and SOCI<NUM>, reaction temperature of about <NUM> and residence time of about <NUM> affording ethyl shikimate (<NUM> %) compared to previously reported reaction times of up to <NUM> hours in batch. Although the reaction is adequate, the use of SOCl<NUM> is undesirable from an environmental, health and safety perspective.

Due to the greenhouse gas (SO<NUM> and HCl) by-products produced, and toxicity concerns associated with SOCI<NUM>, its replacement is desirable from an environmental, health and safety perspective.

In an attempt to develop a greener process, the use of (COCl)<NUM> as the catalyst in shikimic acid esterification was investigated. Shikimic acid <NUM> was esterified with ethanol in the presence of (COCl)<NUM> to afford ethyl shikimate <NUM>. Reaction optimisation was done in a <NUM>µl glass micro reactor system. Shikimic acid (<NUM>) in ethanol was treated with (COCl)<NUM> (<NUM>, effectively <NUM> equiv. ) in a continuous flow system.

There is a general increase in shimikic acid conversion as residence time and temperature increases (<FIG>). However, shikimic acid conversion decreased at <NUM>. For example, there was <NUM> % shikimic acid conversion at <NUM> and <NUM> residence time compared to a surprising conversion of <NUM> % at <NUM> and <NUM> residence time. The optimum conditions for shikimic acid esterification in the presence of (COCl)<NUM> were found to be about <NUM> to about <NUM>, preferably about <NUM> to about <NUM>, most preferably about <NUM>, and at about <NUM> residence time to afford an unexpected conversion of <NUM> %. Under these conditions, (COCl)<NUM> proved to be a better esterification catalyst than SOCl<NUM> (<NUM> %, <NUM> and <NUM> residence time). In addition, the use of (COCl)<NUM> is more desirable since it is safer and more environmentally friendly than SOCl<NUM>.

We also investigated the use of (COCl)<NUM> in the presence of catalytic amount of DMF, which forms a Vilsmeier reagent which is reported to be a better carboxylic acid activator. However, results showed that the DMF catalysed procedure resulted in unexpectedly lower shikimic acid conversion than the uncatalysed reaction.

Esterification of shikimic acid with ethanol in a continuous flow system using benzene sulphonic acid (BSA) was investigated. Shikimic acid (<NUM>, <NUM> equiv. ) in ethanol was treated with BSA (<NUM> equiv. ) in a continuous flow system. The results are shown in <FIG>.

As can be seen from <FIG>, an increase in both residence time and temperature resulted in an increase in shikimic acid conversion, with a conversion plateau at <NUM> %. Furthermore, the use of highly anhydrous ethanol was preferred for this shikimic acid esterification procedure to afford high conversions. The optimum conditions for shikimic acid Fischer esterification using BSA were found to be about <NUM> and about <NUM> residence time affording <NUM> % shikimic acid conversion. Although slightly more efficient, from a health, environmental and safety perspective, BSA may be considered a more desirable esterification catalyst than SOCl<NUM>.

The use of PTSA as a catalyst for shikimic acid esterification with ethanol in a continuous flow system was investigated. Shikimic acid (<NUM>) in ethanol was treated with catalytic amount of PTSA (<NUM> equiv. ) in a continuous flow system to afford ethyl shikimate <NUM>. The results are shown in <FIG>.

In the presence of catalytic amount of PTSA, shikimic acid conversion increased with increase in both residence time and temperature. Better conversions were achieved at <NUM> and above. The optimum conditions for the PTSA catalysed shikimic acid esterification were found to be about <NUM> and about <NUM> residence time to afford <NUM> % conversion. Although better reaction efficiency was observed with SOCI<NUM>, from a health, environmental and safety perspective, PTSA may be considered a more desirable esterification catalyst than SOCl<NUM>.

In order to avoid generating substantial quantities of acid waste, the use of a solid acid catalyst Amberlyst <NUM> in shikimic acid esterification with ethanol in a continuous flow system was investigated (Scheme <NUM>).

In the presence of dry Amberlyst <NUM> as the catalyst for shikimic acid esterification, shikimic acid conversion increased with increase in both temperature and residence time (<FIG>). It appears that the reaction reached a conversion plateau at about <NUM> %. The general recommended operating temperature for Amberlyst <NUM> is <NUM>. However, the inventors unexpectedly found that it was possible to investigate the shikimic acid esterification using Amberyst <NUM> at higher temperatures, including at about <NUM>.

To their delight they observed better conversions at <NUM> (<NUM> % at <NUM> residence time) than at <NUM> (<NUM> % at <NUM> residence time). In the experimental setup, optimum conditions were found to be <NUM> and <NUM> residence time to afford <NUM> % shikimic acid conversion. This compares favourably with the optimum conditions (<NUM> %, <NUM> and <NUM> residence time) found for shikimic acid esterification using SOCl<NUM>. Furthermore, from a health, environmental and safety point of view, the Amberlyst <NUM> procedure is more desirable than the hazardous SOCl<NUM> procedure. As an additional advantage, Amberlyst <NUM> can be removed at the end of the reaction, and regenerated for further use. Amberlyst <NUM> was also evaluated. However, results showed that Amberlyst <NUM> requires almost twice the residence time needed for Amberlyst <NUM> and SOCl<NUM> to afford the same shikimic acid conversion.

Ethyl shikimate <NUM> mesylation was performed in a <NUM> PTFE coil reactor system under sonication (Scheme <NUM>).

A <NUM> PTFE coil reactor (<NUM> ID, <NUM> tube length) under sonication (Scheme <NUM>) was used to optimise the mesylation of ethyl shikimate <NUM> to afford the trimesylate <NUM>. Ethyl shikimate <NUM> (<NUM>) was premixed with mesyl chloride (<NUM>, <NUM> equiv. ) in ethyl acetate to make the first solution. Ethyl shikimate <NUM> is not freely soluble in ethyl acetate. Consequently, it is dissolved in hot ethyl acetate first then cooled before premixing with mesyl chloride. The second solution was made by dissolving an organic base in ethyl acetate. The following bases were screened: triethyl amine (TEA) Imidazole, <NUM>,<NUM>-Diazabicyclo[<NUM>. <NUM>]undec-<NUM>-ene (DBU), <NUM>,<NUM>-diazabicyclo[<NUM>. <NUM>]octane (DABCO) and trihexylamine (THA). The samples collected were first filtered through a PTFE syringe filter (<NUM>µl pore size) to remove the ammonium salts which formed during the reaction before being analysed using HPLC method B.

Ethyl shikimate <NUM> mesylation is reportedly done in batch type reactions using MsCI in the presence triethyl amine (TEA) as the base at low temperatures, preferably <NUM> for about <NUM> to <NUM> hours. Although mesylation transformations in batch are well established and documented, to the best of our knowledge there is no literature on continuous flow ethyl shikimate mesylation, any mesylation in a synthesis method towards Tamiflu, let alone simple model mesylation reactions.

Preliminary attempts for ethyl shikimate (<NUM>) mesylation within a <NUM>µl Chemtrix glass microreactor (<NUM> channel width, <NUM> channel depth) using MsCI (<NUM>, effectively <NUM> equiv. ) and TEA (<NUM> equiv. ) in ethyl acetate failed due to spontaneous blockages. These blockages were due to the ammonium salt precipitate formed between MsCI and TEA during the reaction. Experiments at lower concentrations present the same problem.

In an attempt to investigate a reactor with bigger channel diameter, we used a Little Things Factory (LTF) glass reactor with <NUM> channel diameter. Experiments with a simple PTFE tube coil reactor (<NUM> ID) was also attempted. Disappointingly, none of these attempts solved the reactor blockage issue, even at very low concentrations.

However, in another experiment we employed a <NUM> PTFE coil reactor (<NUM> ID, <NUM> tube length) under sonication (Scheme <NUM>). Ultrasonication appears to have assisted with movement of the ammonium salt precipitate, thereby avoiding reactor blockages. Consequently, this development enabled us to investigate different reaction parameters and ultimately reaction optimisation. It is however envisaged that ultrasonication may not necessarily be required when the reaction is scaled to industrial scale.

Ethyl shikimate b premixed with MsCI in ethyl acetate was treated with TEA in a sonicated continuous flow system affording trimesylate <NUM>. Studies in this system were done using ethyl shikimate (<NUM>), MsCI (<NUM>, effectively <NUM> equiv. ) and TEA (<NUM>, <NUM> equiv. ) in ethyl acetate at <NUM>. The effect of residence time on the reaction is shown in <FIG>.

Ethyl shikimate conversion increased with an increase in residence time (<FIG>). However, it was suprising to note that an increase in residence time did not significantly increase conversion as ethyl shikimate conversion. Although, the reaction is reportedly preferably done at <NUM> in batch, the use of higher temperatures to improve conversions in continuous flow systems were investigated. Room temperature and <NUM> were investigated and there was no conversion improvement. Further investigations were therefore conducted at room temperature.

Since an increase in both residence time and reaction temperature resulted in insignificant ethyl shikimate conversion, the effect of increasing base (TEA) concentration was investigated. Ethyl shikimate (<NUM>, <NUM> equiv. ), MsCI (<NUM> equiv. ) at room temperature and <NUM> residence time was used for these experiments whilst varying TEA concentration. The results of these experiments are shown in <FIG>.

As can be seen from <FIG>, ethyl shikimate conversion increased with an increase in base (TEA) concentration. The optimum conditions were found to be ethyl shikimate (<NUM>), MsCI (<NUM>, effectively <NUM> equiv. ), TEA (<NUM>, <NUM> equiv. ) at room temperature and <NUM> residence time affording the desired mesylate in <NUM> % conversion. The observations indicated that the reaction could be done even at much lower residence times than <NUM>. However, it was difficult to do a comprehensive investigation of lower residence times due to the limitations posed by the syringe pumps available.

The use of bases other than TEA in continuous flow ethyl shikimate mesylation was investigated. A continuous flow system under sonication (Scheme <NUM>) at room temperature and <NUM> residence time was used for these experiments. Ethyl shikimate (<NUM>, <NUM> equiv. ) premixed with MsCI (<NUM> equiv. ) in ethyl acetate was treated with a suitable base (<NUM> equiv. ) in a continuous flow system. The results are shown in <FIG>.

From <FIG>, it can be seen that TEA highest conversion of the bases investigated bases, while DABCO performed the worst. Apart from DABCO, all the investigated bases gave results comparable to TEA. Ammonium salt precipitation remained problematic. However, a lighter precipitate was observed with DBU and imidazole. The use of THA interestingly gave a clear solution. The precipitate absence can be attributed to the increase in hydrophobicity (THA) as chain length increased compared to TEA which made the ammonium salt formed soluble in the reaction solvent ethyl acetate.

Stereoselective and regioselective nucleophilic substitution of the OMs group at the allylic C-<NUM> position by azido group was done by using different azidating agents and conditions (Scheme <NUM>).

Chemtrix's Labtrix start continuous flow system fitted with a <NUM>µl glass reactor was used to optimise the azidation of the OMs group at the allylic C-<NUM> position of mesyl shikimate <NUM> in the presence of various azidating agents (Scheme <NUM>). Sodium azide (NaN<NUM>), diphenylphosphoryl azide (DPPA), trimethylsilyl azide (TMSA) and tetrabutyl ammonium azide (TBAA) were the various azidating agents investigated in this system. The reaction was quenched within the flow reactor using aqueous HCl (<NUM>, <NUM> equiv. ) when necessary. Samples were collected and analysed using HPLC method A.

Mesyl shikimate <NUM> in the presence of a suitable azidating agent undergoes a highly regio- and stereoselective nucleophilic substitution of allylic O-mesylate at the C-<NUM> position affording azide compound <NUM> (Scheme <NUM>).

Initial experiments had shown the same conversions in both acetone and acetonitrile as mesyl shikimate solvents. However, the use of acetone was associated with eventual micro reactor blockage caused by a resulting precipitate from acetone-aqueous NaN<NUM> mixture. Furthermore, acetonitrile has a higher boiling point than acetone which is desirable for high temperature reaction interrogation. Consequently, acetonitrile was the preferred solvent for mesyl shikimate <NUM> for further optimisation in continuous flow systems.

Mesyl shikimate <NUM> (<NUM>) in acetonitrile was treated with aqueous NaN<NUM> (<NUM>, <NUM> equiv. ) in a thermally controlled micro reactor system (Scheme <NUM>). The reaction generally affords two products, the desired azide compound <NUM> and a side product 41a (Scheme <NUM>).

The findings on the effect of various reaction conditions on conversion and selectivity are shown in <FIG> and <FIG>.

From <FIG>, it is evident that mesyl shikimate conversion increases with increase in residence time and temperature. At <NUM> and above, full conversions were achieved at low residence times. Full conversion was achieved at <NUM>, <NUM> residence time and <NUM> % conversion at <NUM> ° C and <NUM> residence time (<FIG>).

Product selectivity to azide <NUM> is shown in <FIG>. As can be seen from <FIG>, selectivity generally decreases with increase in residence time and temperature. However, there is <NUM> % selectivity towards the desired azide <NUM> at <NUM> at all the investigated residence times. At <NUM>, <NUM> % and <NUM> % azide <NUM> selectivity was obtained at <NUM> and <NUM> respectively. It is evident that high temperatures favour the undesired aromatic azide compound 41a.

The effect of NaN<NUM> concentration on the reaction was investigated, thereby to determine the effect of NaN<NUM> molar equivalent on mesyl shikimate <NUM> conversion and selectivity of the desired azide <NUM> at <NUM> and <NUM> residence time. Results showed that selectivity towards azide <NUM> decreases with increase in NaN<NUM> concentration. Contrary, mesyl shikimate <NUM> conversion improved with an increase in NaN<NUM> (<NUM> and <NUM> equivalents). It is believed that the excess NaN<NUM> increases reaction basicity, resulting in the undesired azide 41a being exceedingly favoured.

The preferred conditions in flow for this reaction were found to be <NUM> equivalents of NaN<NUM>, <NUM> and <NUM> residence time affording full conversion to towards the desired azide <NUM>. Despite of high temperatures, long reaction times and basicity being detrimental to the selectivity of the desired azide <NUM> in batch, as previously reported, it is evident from our experiments that microreactor use significantly improved selectivity let alone massively reducing reaction times.

Alternative, non-aqueous, azidation procedures in continuous flow systems were also investigated. Azidating agents investigated include diphenyl phosphoryl azide (DPPA), trimethylsilyl azide (TMSA), and tetrabutylammonium azide (TBAA).

Mesyl shikimate (<NUM>) was treated with a mixture of DPPA (<NUM>, <NUM> equiv. ) and TEA (<NUM>, <NUM> equiv. ) in a continuous flow system. The reaction was quenched with aqueous HCl (<NUM>, <NUM> equiv. ) within the flow system. Conversion and selectivity results for this reaction is shown in <FIG> and <FIG>.

As can be seen from <FIG>, with DPPA as the azidating agent an increase in both temperature and residence time resulted in the increase in mesyl shikimate conversion in microreactors. Azide <NUM> selectivity decreases with increase in temperature and residence time (<FIG>). The trends observed in respect of DPPA were similar to those observed with NaN<NUM> as the azidating agent.

The lower azide <NUM> selectivity associated with DPPA is as result of the base used. Basic conditions are reportedly detrimental to the azide <NUM> selectivity. The use of a base in the DPPA procedure was unavoidable as the reaction did not proceed in its absence. The reaction was quenched within the microreactor using aqueous HCl. The effect of base (TEA) concentration on azide <NUM> selectivity was investigated at room temperature and <NUM> residence time to ascertain its role in the formation of the unwanted aromatic azide 41a. Results showed that selectivity towards the desired azide <NUM> significantly decreased with an increase in base concentration.

The use of TMSA as the azidating agent was also investigated. The conversions found with TMSA were comparable with both NaN<NUM> and DPPA. Mesyl shikimate conversion of <NUM> %, <NUM> % and <NUM> % were obtained at <NUM> and <NUM> residence time by using TMSA, DPPA and NaN3 as azidating agents respectively. Azide <NUM> selectivity using TMSA was also comparable to that obtained with DPPA and NaN<NUM>.

Reactions performed with TBAA resulted in unacceptable selectivity towards azide <NUM>, while the use of azide ion exchange reson (Amberlite IRN78) gave comparatively poor conversion performance.

The preferred reaction conditions for the NaN<NUM> procedure were found to be about <NUM> equivalents of NaN<NUM>, <NUM> and <NUM> affording full conversion towards the desired azide <NUM>. Contrary to all the published literature procedures, side product 41a was not produced using our procedure. The preferred conditions for the anhydrous procedure are about <NUM> equivalents of DPPA/TMSA, about <NUM> equivalents of TEA, <NUM> and <NUM> affording about <NUM> % conversion towards the desired azide <NUM>.

Continuous flow aziridination of azide <NUM> was done in a Chemtrix Labtrix flow system (Scheme <NUM>).

Chemtrix's Labtrix start continuous flow system fitted with a <NUM>µl glass reactor was used to optimise the aziridination of the azido shikimate <NUM> using trialkyl phosphite. Triethyl phosphite and trimethyl phosphate were the two alkyl phosphite investigated. A solution of azido shikimate in anhydrous acetonitrile (<NUM>) and a solution of trialkyl phosphite in anhydrous acetonitrile (<NUM>, <NUM> equiv. ) were pumped separately using two syringe pumps from two <NUM> SGE Luer lock gas tight glass syringes into the thermally controlled microreactor system which was fitted with a <NUM> bar (<NUM><NUM> kPa) back pressure regulator (Scheme <NUM>). Samples were collected and analysed using HPLC method A.

The azide <NUM> undergoes aziridination in the presence of trialkyl phosphite under water-free conditions (Scheme <NUM>). In one set of experiments, azide <NUM> was treated with (EtO)<NUM>P under anhydrous conditions in a continuous flow system to afford azidine <NUM>.

Azide <NUM> (<NUM>) was treated with (EtO)<NUM>P (<NUM>, <NUM> equiv. ) and spontaneously reacted affording full conversion towards an undesirable product. This was suggestive of an intermediate formation. Experiments were then conducted with acetonitrile as solvent, with the only possible disadvantage of acetonitrile being its lower boiling point, compared to toluene. This was compensated for by pressurising the system, which allowed for reaction superheating without solvent boiling.

A solution of azide <NUM> (<NUM>) in acetonitrile was treated with (EtO)<NUM>P (<NUM>, <NUM> equiv. ) to afford aziridine <NUM>. The conversion of azide <NUM> to aziridine <NUM> at various conditions in flow is shown in <FIG>. Conversion of azide <NUM> to aziridine <NUM> in the presence of (EtO)<NUM>P increased with increase in temperature and residence time. Good conversion was achieved at very fast residence times (< <NUM>). Temperature was found to have the most significant effect on the successful synthesis of aziridine <NUM>. As mentioned above, azide <NUM> was completely consumed in the presence of (EtO)<NUM>P at room temperature (toluene), however no aziridine <NUM> was detected. The use of pressurised microreactors allowed for very high reaction temperatures which resulted in very fast reactions. Furthermore, microreactors allowed safe interrogation of potentially explosive azide chemistry at very high temperatures. Optimum conditions were found to be about <NUM> and <NUM> residence time to afford full conversion towards the desired aziridine <NUM>.

In another set of experiments, a continuous flow system was used to perform azide <NUM> aziridination reaction with (MeO)<NUM>P to afford aziridine <NUM>. Results showed that, as with (EtO)<NUM>P, aziridine formation increased with increased temperature and residence time. The use of (MeO)<NUM>P proved to be slightly more efficient in aziridination than (EtO)<NUM>P. At about <NUM> and <NUM> residence time, <NUM> % and <NUM> % aziridine <NUM> was formed using (EtO)<NUM>P and (MeO)<NUM>P respectively.

Importantly, our system and process allowed for high temperature azide chemistry, resulting in very fast reactions compared to the <NUM> hour batch reactions previously reported.

The aziridine <NUM> underwent regio- and stereoselective ring opening with <NUM>-pentanol and the Lewis catalyst boron trifloride etherate at the allylic position (Scheme <NUM>) in a continuous flow system.

Chemtrix's Labtrix start continuous flow system fitted with a <NUM>µl glass reactor was used to optimise the aziridine <NUM> ring opening with <NUM>-pentanol and boron trifloride etherate (Scheme <NUM>). The aziridine <NUM> (<NUM>) in acetonitrile/<NUM>-pentanol (<NUM>:<NUM>) and a solution of boron trifloride etherate (<NUM>, <NUM> equiv. ) in acetonitrile/<NUM>-pentanol (<NUM>:<NUM>) were pumped separately using two syringe pumps from two <NUM> SGE Luer lock gas tight glass syringes into the thermally controlled microreactor system which was fitted with a <NUM> bar (<NUM><NUM> kPa) back pressure regulator. Samples were collected and analysed using HPLC method A.

Initial experiments were performed by treating aziridine <NUM> (<NUM>) in <NUM>-pentanol with BF<NUM>. OEt<NUM> (<NUM>, <NUM> equiv. ) in <NUM>-pentanol at <NUM> in a continuous flow system for <NUM> residence time affording <NUM>-pentyl ether <NUM> in <NUM> % conversion. The use of an excess of BF<NUM>. OEt<NUM> (<NUM> equiv. ) resulted in full conversion towards <NUM>-pentyl ether <NUM>. Due to pressure build up in the system at higher flow rates, attributed to high viscosity of <NUM>-pentanol, residence times lower than <NUM> could not be investigated in this system. Therefore, preliminary investigations at residence times less than <NUM> were successfully done by using a <NUM>-pentanol/acetonitrile (<NUM>/<NUM>) mixture.

The reaction was optimised in a continuous flow system by treating aziridine <NUM> (<NUM>) in <NUM>-pentanol/acetonitrile (<NUM>/<NUM>) with BF<NUM>. OEt<NUM> (<NUM>, <NUM> equiv. ) in <NUM>-pentanol/acetonitrile (<NUM>/<NUM>) at various reaction conditions. The use of diluted <NUM>-pentanol allowed us to interrogate the reaction at very fast reaction times. Results of these experiments are shown in <FIG>.

As can be seen from <FIG>, the conversion towards <NUM>-pentyl ether <NUM> from aziridine <NUM> increased with increase with residence time and temperature. Temperature increase resulted in a significant improvement in conversion. At <NUM> residence time, a <NUM>-pentyl ether <NUM> yield of <NUM> % and <NUM> % was achieved at <NUM> and <NUM> respectively. The preferred conditions were found to be about <NUM> and <NUM> residence time to afford full conversion towards <NUM>-pentyl ether <NUM>.

Acetylation of the <NUM>-pentyl ether <NUM> was achieved via N-P bond cleavage with sulphuric acid, followed by acetylation under slightly basic conditions (Scheme <NUM>). <CHM>
Intermediate 43a was formed in situ in the first thermally controlled reactor by treating the <NUM>-pentyl ether <NUM> (<NUM>) in acetonitrile with H<NUM>SO<NUM> (<NUM>, <NUM> equiv. ) in acetonitrile. The intermediate 43a formed in situ was treated with NaOH (<NUM>, <NUM> equiv. ) and then acetic anhydride (<NUM> equiv. ) in the second thermally controlled reactor to afford acetamide <NUM>. This system for multistep continuous flow system was fitted with a <NUM> bar (<NUM><NUM> kPa) back pressure regulator. Samples were collected and analysed using HPLC method A.

In a preliminary set of experiments, <NUM>-pentyl ether <NUM> (<NUM>) in ethanol was treated with H<NUM>SO<NUM> (<NUM>, <NUM> equiv. ) in ethanol at <NUM> and <NUM> residence time afforded compound 43a in <NUM> % conversion. Comparable results were obtained when acetonitrile was used as a solvent instead of ethanol. Since acetonitrile was the common solvent for most of the continuous flow steps, the reaction was then optimised in acetonitrile. A solution of <NUM>-pentyl ether <NUM> (<NUM>) in acetonitrile was treated with a solution of H<NUM>SO<NUM> (<NUM>, <NUM> equiv. ) in acetonitrile in a Chemtrix microreactor at various reaction conditions for optimisation. The results of these experiments are shown in <FIG>.

The conversion of <NUM>-pentyl ether <NUM> to compound 43a increased with increase in temperature and residence time. Conversion towards compound 43a was <NUM> % and <NUM> % at <NUM> and <NUM> respectively at <NUM> residence time. At constant temperature of <NUM>, conversion towards compound 43a was <NUM> % and <NUM> % at <NUM> and <NUM> respectively. The preferred conditions for continuous flow N-P bond cleavage were found to be about <NUM> and <NUM> residence time using H<NUM>SO<NUM> (<NUM> equiv. ) to afford compound 43a in full conversion.

Having successfully demonstrated the continuous flow N-P bond cleavage in <NUM>-pentyl ether <NUM> afford compound 43a, the investigation was extended to acetylation of compound 43a.

The acetylation of <NUM>-pentyl ether <NUM> was completed by subsequently treating the compound 43a formed in situ with Ac<NUM>O under slightly basic conditions to afford acetamide <NUM> in a continuous flow system (Scheme <NUM>). Based on results from previous experiments, <NUM>-pentyl ether <NUM> (<NUM>) in acetonitrile was treated with H<NUM>SO<NUM> (<NUM> equiv. ) in acetonitrile at <NUM> in the first microreactor for <NUM> residence time to afford compound 43a in situ which was subsequently treated with an aqueous NaOH (<NUM>, <NUM> equiv. ) and subsequently with Ac<NUM>O (<NUM>, <NUM> equiv. ) at room temperature to afford acetamide <NUM> (<NUM> %) at <NUM> total residence time.

Increase in total residence time for acetylation resulted in increase in acetamide <NUM> formation. Doubling the total residence time resulted in a <NUM> % conversion increase. Total residence time of <NUM> gave <NUM> % acetamide <NUM> and there was no conversion improvement beyond <NUM> residence time. As in the preceding steps, basic reaction medium could have caused side reactions thus lowering acetamide <NUM> yield. The basic reaction medium is attributed to the slightly basic medium used to facilitate acetylation and the basicity of intermediate 43a formed in situ.

Acetamide <NUM> is treated with a suitable azidating agent to afford azide <NUM>. The C-<NUM> OMs group on acetamide <NUM> undergoes nucleophilic replacement by the N<NUM> group (Scheme <NUM>).

Chemtrix's Labtrix start continuous flow system fitted with a <NUM>µl glass reactor was used to optimise the C-<NUM> azidation of acetamide <NUM> (Scheme <NUM>). The acetamide <NUM> (<NUM>) in acetonitrile and azidating agent in appropriate solvent (<NUM>, <NUM> equiv. ) were pumped separately using two syringe pumps from two <NUM> SGE Luer lock gas tight glass syringes into the thermally controlled microreactor system which was fitted with a <NUM> bar (<NUM><NUM> kPa) back pressure regulator. The use of NaN<NUM>, TBAA, DPPA, and TMSA were investigated. Samples were collected and analysed using HPLC method A.

Initial experiments in flow were done using acetamide <NUM> (<NUM>) in DMF and aqueous NaN<NUM> (<NUM>, <NUM> equiv. ) in a <NUM>µl glass microreactor at <NUM> for <NUM> affording azide <NUM> (<NUM> %). In other experiments we achieved <NUM> % conversion to azide <NUM> in acetonitrile. The reaction was further optimised using acetonitrile as acetamide <NUM> (<NUM>) solvent and aqueous NaN<NUM> (<NUM>, <NUM> equiv. The results of these experiments are shown in <FIG>.

As can be seen from <FIG>, azide <NUM> formation is a function of temperature and residence time. The conversion of acetamide <NUM> to azide <NUM> increased with increase temperature. Conversion towards azide <NUM> was <NUM> % and <NUM> % at <NUM> and <NUM> at <NUM> residence time respectively. The preferred conditions were found to be about <NUM>, <NUM> residence time to afford azide <NUM> in full conversion.

Preferred conditions developed for NaN<NUM> (<NUM> equiv. , <NUM>, and <NUM>) were used to investigate the use DPPA, TMSA and TBAA as azidating agents for acetamide <NUM> in a <NUM>µl glass microreactor. In these experiments, acetamide <NUM> was successfully converted to azide <NUM> at varying conversions DPPA, TMSA, and TBAA. It appears that the application of ionic bonded azides (NaN<NUM> and TBAA) gave similar conversions (<NUM> % and <NUM> % respectively), whilst covalently bonded azides (DPPA and TMSA) resulted in comparativelt lower conversions (<NUM> % and <NUM> % respectively).

A <NUM> PTFE coil reactor (<NUM> ID, <NUM> tube length) under sonication (Scheme <NUM>) was used to optimise the azide <NUM> reduction to afford Oseltamivir using NaBH<NUM> and CoCl<NUM>. A mixture of azide <NUM> (<NUM>) with CoCl<NUM> (<NUM> equiv. ) in ethanol and NaBH<NUM> (<NUM>, <NUM> equiv. ) in water (pH = <NUM>) was pumped through the continuous flow system to afford Oseltamivir <NUM>. The samples collected were first filtered through a PTFE syringe filter (<NUM>µl pore size) to remove the cobalt boride precipitates formed in the reaction, and then analysed using HPLC method A.

Oseltamivir <NUM> was synthesised from CoCl<NUM> catalysed NaBH<NUM> reduction of azide <NUM> in a continuous flow system under sonication because the formation of a black precipitate (cobalt boride) was observed in preliminary batch-type investigations. However, it is possible that the formation of this precipitate may not be problematic when the method is performed using industrial scale equipment.

A mixture of azide <NUM> (<NUM>) with CoCl<NUM> (<NUM> equiv. ) in ethanol was treated with NaBH<NUM> (<NUM>, <NUM> equiv. ) in buffered water (pH = <NUM>) in a <NUM> PTFE coil reactor system (<NUM> ID, <NUM> tube length) under sonication to afford oseltamivir <NUM>. The results of these experiments are shown in <FIG>. Conversion towards Oseltamivir <NUM> increased with increase in residence time. Conversion towards oseltamivir <NUM> was unexpectedly found to be <NUM> % and <NUM> % at residence times of only <NUM> and <NUM> respectively. The preferred conditions were found to be about room temperature and a residence time of about <NUM> to afford Oseltamivir <NUM> (<NUM> %).

We also conducted experiments to investigate the use of ethanol and methanol for NaBH<NUM>. These experiments were done at room temperature for <NUM> residence time in a sonicated continuous flow system. Although ethanol and methanol are known to react with NaBH<NUM>, it appears that the azide reduction reaction, in the presence of a catalytic amount of CoCl<NUM>, is so fast that it overshadows the competing reactions by these solvents. Therefore, although water at ph > <NUM> is preferred, it is anticipated that several other solvents will also be suitable.

In other experiments, as suggested in known batch methods, a phosphine based reaction was attempted in flow by treating azide <NUM> (<NUM>) in THF with Ph<NUM>P (<NUM>, <NUM> equiv. ) in THF/water (<NUM>:<NUM>) in continuous flow system at <NUM> to afford Oseltamivir <NUM>. The conversion of azide <NUM> towards Oseltamivir <NUM> was <NUM> % and <NUM> % at <NUM> and <NUM> residence time respectively, which is much lower than that obtained in the preferred reaction with CoCl<NUM> and NaBH<NUM>.

Oseltamivir <NUM> was treated with H<NUM>PO<NUM> to afford Oseltamivir phosphate <NUM> (Scheme <NUM>) in a continuous flow system.

A <NUM> PTFE coil reactor (<NUM> ID, <NUM> tube length) under sonication was used to optimise the treatment of Oseltamivir <NUM> with H<NUM>PO<NUM> affording Oseltamivir phosphate <NUM> in a continuous flow system. Oseltamivir <NUM> (<NUM>) in ethanol and H<NUM>PO<NUM> (<NUM>, <NUM> equiv. ) in ethanol were pumped through a thermally controlled continuous flow system to afford oseltamivir phosphate. Samples were collected and analysed using HPLC method A.

Oseltamivir <NUM> (<NUM>) in ethanol was treated with H<NUM>PO<NUM> (<NUM>, <NUM> equiv. ) in ethanol at <NUM> in a <NUM> PTFE coil reactor (<NUM> ID, <NUM> tube length) under sonication at various residence times for optimisation. The results of these experiments are shown in <FIG>.

Claim 1:
A flow synthesis process for producing a compound of the Formula <NUM> and its pharmaceutically acceptable salts,
<CHM>
the process comprising the steps of:
a) preparing ethyl shikimate of Formula <NUM>
<CHM>
by reacting shikimic acid with a reagent selected from the group consisting of (COCI)<NUM>, SOCI<NUM>, benzene sulphonic acid (BSA), p-toluene sulphonic acid (PTSA), and Amberlyst <NUM>,
b) reacting the ethyl shikimate of Formula <NUM> with mesyl chloride in the presence of a base selected from the group consisting of trimethyl amine (TEA), <NUM>,<NUM>-diazabicyclo(<NUM>.<NUM>)undec-<NUM>-ene (DBU), imidazole, and trihexyl amine (THA) to produce the O-trimesylate of Formula <NUM>
<CHM>
c) reacting the O-trimesylate of Formula <NUM> in an azidation reaction with an appropriate azidating agent to produce the azide of Formula <NUM>
<CHM>
d) reacting the azide of Formula <NUM> in an aziridation reaction with a trialkyl phosphite to produce the aziridine of Formula <NUM>
<CHM>
e) reacting the aziridine of Formula <NUM> with <NUM>-pentanol in the presence of a Lewis acid catalyst to produce the <NUM>-pentyl ether of Formula <NUM>
<CHM>
f) reacting the <NUM>-pentyl ether of Formula <NUM> in an acetylation reaction with H<NUM>SO<NUM>, followed by Ac<NUM>O in the presence of a suitable base to produce the acetamide of Formula <NUM>
<CHM>
,
g) reacting the acetamide of Formula <NUM> in an azidation reaction with an appropriate azidating reagent produce the azide of Formula <NUM>
<CHM>
and
h) reacting the azide of Formula <NUM> by mixing the azide with CoCl<NUM>, and reacting the mixture with NaBH<NUM> to produce the compound of Formula <NUM>.