Title of Invention

PROCESS FOR SEPARATING A SINGLE STEREOISOMER FROM A RACEMIC MIXTURE

Abstract

159/CHENP/2003 "Process for separating a single stereoisomer from a racemic mixture" This invention relates to a process of separating a single desired stereoisomer from a racemic mixture of eight stereoisomers of a compound of formula (III), wherein R1 represents an isopropanol group, an isopropyl group or an isopropylene group, includes the steps of contacting the racemic mixture in a suitable organic solvent with an esterifying agent and a stereospecific enzyme which stereoselectively esterifies the -OH group of the desired stereoisomer, for a time sufficient to convert a desired percentage of the desired stereoisomer to a compound of formula (IV), wherein R1 is as defined above and R4 is an alkyl or an aryl group, to give a first reaction product including the compound of formula (IV), the organic solvent, the unconverted stereoisomers of the compound of formula (III), excess esterifying agent and by-products of the reaction; and separating the compound of formula (IV) from the first reaction product. The process is of particular application for the production of (-)-menthol.

Full Text

This invention relates to a process of separating a single stereoisomer from a racemic mixture. (-)-Menthol is one of the world's largest selling flavour compounds with a production of about 11800 tons per annum. Its peppermint flavour and cooling sensation are used in many products, primarily m mentholated cigarettes and oral hygiene products, such as toothpaste, mouthwash and chewing gum. Pharmaceutical and healthcare products use menthol in a variety of product types, such as cough lozenges, shaving cream and topical analgesics. Owing to seasonal variations and poor farming practices, the availability of natural menthol from the largest supplying countries, India and China is sometimes erratic. In addition to this, only a limited amount of peppermint can be planted, thus limiting the supply of natural menthol. The remainder of the demand for menthol is met by synthetic menthol. Organoleptically, natural and synthetic menthol are completely interchangeable; any slight differences have been largely eliminated through product development. Moreover, synthetic menthol has a purer and more consistent taste and odour profile since it does not contain the impurities present in natural menthol. There is always a need for new processes for the production of (-)-menthol and similar compounds. SUMMARY OF THE INVENTION According to the invention there is provided a process of separating a single desired stereoisomer from a racemic mixture of eight stereoisomers of a compound of the formula 111 Wherein R1 represents an isopropanol group, an isopropyl group or an isopropylene group, including the steps of: (1) contacting the racemic mixture in a suitable organic solvent with an esterifying agent and a stereospecific enzyme which stereoselectively esterifies the -OH group of the desired stereoisomer, for a time sufficient to convert a desired percentage of the desired stereoisomer to a compound of the formula IV I wherein R1 is as defined above and R4 is an alkyl or an aryl group, to give a first reaction product including the compound of the formula IV. the organic solvent, the unconverted stereoisomers of the compound of the formula ill, excess esterifying agent and by-products of the reaction; and (2) separating the compound of the formula IV from the first reaction product. Step (2) preferably comprises the sub-steps of: (2)(a) separating the first reaction product from the enzyme; (2)(b) removing from the first reaction product the organic solvent, the excess esterifying agent, and the by-products of the reaction to give a second reaction product including the compound of the formula IV.and the unconverted stereoisomers of the compound of formula 111; and (2)(c) separating the compound of the formula iV from the second reaction product leaving a third reaction product containing the unconverted stereoisomers of the compound of the fomiula III, The process of the invention preferably includes a further step, step (3) of: (3) racemizing the unconverted stereoisomers of the compound of the fomnula III in the third reaction product to give a fourth reaction product containing a mixture of all eight stereoisomers of the compound of the formula III and recycling the fourth reaction product to step (1). The process of the invention preferably includes a further step, step (4) of: (4) hydrolysing the compound of the formula IV to give the desired stereoisomer of the compound of the formula 111. In the process of the invention, when R1is an isopropanol group or an isopropylene group, before or after step (4), the compound of the formula IV or the desired stereoisomer of the compound of the formula 111 may be subjected to a reduction step to convert R1 to an isopropyl group. The process of the invention preferably includes the following steps, prior to step (1) of: (a) reacting a compound of the formula I wherein R2 represents a methyl or a hydroxymethyl group, and R3 represents H, a base metal, a benzyl group or an allyl group, with an alkylating agent in the presence of a catalyst to give a compound of the formula II i wherein R1, R2 and R3 are as defined above; and (b) hydrogenating the compound of the formula II in the presence of a catalyst to give a racemic mixture of the eight stereoisomers of the compound of the formula 111. DESCRIPTION OF EMBODIMENTS The curx of the invention is a process of separating a single desired stereoisomer of a compound of the formula ill from a racemic mixture of the eight stereoisomers of the compound of the formula 111, by esterification using a stereospecific enzyme. This process step may form part of a larger process as described below. The first step, step (a) of the process is to alkylate a compound of the formula I to yield a compound of the formula II, using an alkylating agent such as propylene, isopropanol or acetone, in the presence of a catalyst. The catalyst may be a Lewis acid catalyst such as AlCl3, SnCU, BF3, ZnCl2 or FeCl3; or a Brcnsted acid such as HCI, HF, H2SO4 or H3PO4; or a suitable supported catalyst such as Envirocat EPIC which is a polyphosphoric acid on a support, or Envirocat EPZ 10 which is a ferric chloride on a support; or a solid phosphoric acid; or a suitable zeolite catalyst which is the preferred catalyst. Step (a) may be carried out in the presence of a solvent such as a chlorinated solvent, e.g dichioromethane, chloroform or dichioroethane, but the presence of a solvent is not essential. Step (a) may be carried out as a batch reaction or as a continuous gas or liquid phase reaction. In step (a), the reaction temperature during and after the reaction may be any temperature below 450'*C. Step (a) may be carried out in air or under an inert atmosphere such as argon or nitrogen. The compound of the formula I is preferably m-cresol, and thus the compound of the fonnuia 11 is preferably thymol. At the end of step (a), the catalyst is removed or deactivated, and any solvent present is removed. The required compound of the formula II is isolated from the reaction mixture, for example by distillation. The unreacted alkylating agent and reaction by-products may be recycled to step (a). The second step, step (b) of the process is. to reduce the compound of the formula II to a racemic mixture of the eight stereoisomers of the compound of the fonmula III, by hydrogenation using hydrogen gas over a suitable catalyst. The catalyst may be any catalyst typically used in homogeneous catalytic hydrogenations such as Pd(OAc)2 or in heterogenous catalytic hydrogenations such-as supported palladium, platinum, rhodium, ruthenium, nickel, sponge nickel and 2CuOCr2O3. The preferred catalyst is a nickel catalyst. The catalyst loading may be from 0,01 to 20%, preferably from 0,5 to 5%. Step (b) may be carried out with or without a solvent. If a solvent is used, it may be any solvent typically used for catalytic hydrogenation, for example a hydrocarbon or aqueous caustic. Step (b) is preferably carried out at an elevated temperature of from 80'C to 300°C inclusive, preferably from 160°C to 200°C inclusive. Step (b) is carried out using a hydrogen pressure which is below 50 bar, preferably between 5 and 35 bar inclusive. When the compound of the formula II is thymol, there is formed a racemic mixture of the four pairs of diastereomeric isomers of menthol, viz (±)-menthol, (±)-isomenthol, {+)-neomenthol and (±)-neoisomenthol, as well as two intermediates (±)"menthone and (±)-isomenthone. At the end of step (b), the catalyst is removed, e.g by filtration, or is deactivated, and any solvent present is removed. The next step of the process of the invention, step (1), is the key step of the process. The racemic mixture of the eight stereoisomers of the compound of the formula II) from the hydrogenation step (step (b)) is contacted in a suitable organic solvent with an esterifying agent and with a stereospecific enzyme which stereoselectively esterifies the -OH group of the desired stereoisomer of the compound of the formula III, for a time sufficient to convert a desired percentage of the desired stereoisomer to a compound of the formula IV, to give a first reaction product including the compound of the formula IV, the organic solvent, the unconverted stereoisomers of the compound of the formula III, excess esterifying agent, and by-products of the reaction. The reaction is a stereoselective esterification of a desired stereoisomer of the compound of the formula 111, with the other stereoisomers of the compound of the formula ill remaining substantially unchanged, although small amounts of the other stereoisomers may also be esterified by the enzyme. When R1 of the compound of the formula 111 is an isopropyl group, the compound corresponds to menthol which consists of eight stereoisomers. The desired stereoisomer of menthol is the (-)-isomer, which can be selectively esterified using a suitable enzyme to the {-)-menthyl ester. The suitable organic solvent may be any solvent typically used for enzyme catalysed esterification reactions, including isooctane; n-heptane; decane; methyl cyclohexane; t-butyl methyl ether; xylene; kerosene (C5-C5 paraffins, kerasol 60/115), (C7-C8 paraffins, kerasol 94/125); pentane; cyclohexane; hexane; benzene; butanol; toluene; isopropanol; ethyl lactate; and acetone. The preferred organic solvent is n-heptane. The amount of the organic solvent used relative to the racemic mixture of the compound of the fomnula 111 is preferably in the range of from 5% to 80% racemic mixture to 95% to 20% organic solvent on a volume basis. When the organic solvent is n-heptane, the racemic mixture is preferably used in an amount of 20% v/v in the n-heptane. The esterifying agent may be any suitable esterifying agent such as for example vinyl acetate, butyl acetate, octanoic acid, isopropenyl acetate, vinyl butyrate, ethyl lactate and ethyl acetate. The preferred esterifying agent is vinyl acetate. The esterifying agent may be used in an molar ratio to the desired stereoisomer of the compound of the formula III of 0,5:1 up to 30:1, The prefen'ed molar ratio of the esterifying agent to the desired stereoisomer of the compound of the formula III is about 2:1 when vinyl acetate is used as the esterifying agent. The enzyme is a stereospecific enzyme which stereoselective esterifies the -OH group of the compound of the formula III. The enzyme may be contained within a microorganism, or secreted into a medium required for microorganism growth, or may be available commercially in semi-purified or purified form. Microorganisms can also be provided with the ability to produce a suitable enzyme through the process of genetic engineering of the microorganism. Examples of enzymes and microorganisms that are capable of performing this esterifying process include those exhibiting lipase, esterase or protease-like activity. Suitable enzymes include, but are not limited to: Enzymes supplied by Fiuka: Candida cyfindracea lipase, lipase Hog pancreas, lipase Pseudomonas fluorescens, lipase Aspergillus oryzae, lipase Rhizopus niveus, lipase Rhizomucor miehei, lipase Candida antarctica, lipase Mucor javanicus, lipase Rhizopus arhizus, lipase Peniciliium roqueforti, lipase Candida lipolytica, lipoprotein lipase Pseudomonas sp., type B, lipoprotein lipase Pseudomonas cepacia, lipoprotein lipase Chromobacterium viscosum, esterase Bacillus thermoglucosidasius. esterase Bacillus stearothermophiius, esterase Mucor miehei, esterase hog liver; Enzymes supplied by Altus: Candida rugosa lipase, lipase Mucor. miehei, Candida antarcitica B lipase, Candida antarctica A lipase, Chiro-CLEC-CR, Chiro-CLEC-CR (slurry), porcine liver esterase, Penicillin acyjase, Subtilisin Carlsberg, Chiro-CLEC-BL (slurry), Chiro-CLEC-PC (slurry), Chiro-CLEC-EC (slurry), Aspergillus oryzae protease, PeptiCLEC-TR (slurry); Enzymes supplied by Recombinant Biocatalysis: ESL-001-07, ESL-001-01, ESL-001-01 with stabiliser, ESL-001-02, ESL-001-03, ESL-001-05; Enzymes supplied by Boehringer-Mannheim: Chirazyme L4 (Pseudomonas sp.), Chirazyme L5 (Candida antarctica fraction A), Chirazyme L1 (Burkholderia), Chirazyme L6 (Porcine pancreas), Chirazyme L7, Chirazyme L8; Enzymes supplied by Gist-Brocades: Naproxen esterase, Lipomax, Genzyme, Lipoprotein lipase; Enzymes supplied by Novo: Novozyme 868, Novozyme 435, immobilised Candida antarctica lipase, Nagase enzyme, Lipase A-10FG (Rhizopus javanicus); Enzymes supplied by Amano: Amano AYS, Amano PS, Amano PSD, Amano AKD11,Amano AKD111. The preferred enzyme is Amano AK lipase enzyme supplied by Amano of Japan. The enzyme may be used either in the free form or immobilized on a suitable support which may be diatomaceous earth. The enzyme is preferably used In an amount of from Ig/I to 60g/! of the reaction mixture, i.e the racemic mixture of the eight stereoisomers of the compound of the formula III, the suitable organic solvent and the esterlfying agent. The resolution step is preferably carried out at a temperature of from 20°C to 100*C inclusive and at atmospheric or higher pressure. When the enzyme is Amano AK, the preferred reaction temperature is about 50oC. The resolution reaction is continued for a time sufficient to convert a desired percentage of the desired stereoisomer of the compound of the formula III to the compound of the formula IV. Generally, it is desirable that as much as possible of the desired stereoisomer is converted to the compound of the formula IV without the reaction proceeding to the esterification of the other stereoisomers present in the racemic mixture. The reaction time is preferably about 24 hours or less when the reaction is performed in batch mode. The next step, step (2)(a) of the process of the invention is to separate the first reaction product from the enzyme so that the enzyme can be recycled. This may be achieved for example by centrifugation or by filtration. A major advantage of the process of the invention is that it is possible to recycle the enzyme a number of times to the resolution step, so as to improve the economics of the process. The necessity for enzyme recycle may be eliminated by use of the enzyme in a continuous system wherein the enzyme is retained within a reactor. The racemic mixture of the compound of the formula 111, the organic solvent and the esterifying agent as described above, are fed into the reactor, wherein the desired stereoisomer of the compound of the formula III is esterlfied to the compound of the formula IV to form the first reaction product The first reaction product typically exits the reactor at a similar rate to the inlet feed, for further processing. The enzyme may typically be retained within the reactor through the use of membranes, or by immobilisation onto a support material, or through stabilisation by cross-finking. The next step, step (2)(b) of the process of the invention is to remove the organic solvent, the excess esterifying agent and any by-products, to give a second reaction product including the compound of the formula IV and the unconverted stereoisomers of the compound of the formula III. This may be carried out by distillation in which the organic solvent, e.g the n-heptane and the excess esterifying agent, e.g the vinyl acetate, are taken off at the top of the column as a single stream and recycled back to suitable storage tanks for later re-use. The next step, step (2)(c) of the process of the invention is to separate the compound of formula IV from the second reaction product leaving a third reaction product containing the unconverted stereoisomers of the compound of the formula III. This separation may be achieved by distillation. In the compound of the formula IV, when R1 is not an isopropyl group, the R1 group can, through a reduction process, be converted to an isopropyl group, either at this stage, or subsequent to hydrolysis of the ester group as described below. This results in the production of the desired stereoisomer of menthyl ester, or subsequent to hydrolysis, in the production of the desired stereoisomer of menthol. The next step, step (3) of the process of the invention is to racemise the unconverted stereoisomers in the third reaction product to give a fourth reaction product containing a mixture of all eight stereoisomers of the compound of the formula III and recycling this to the resolution step of the process. The racemisation may be achieved over a suitable catalyst with or without hydrogen gas, with or without a solvent, and at atmospheric or greater pressure. The step may be carried out with or without a solvent. If a solvent is used, the solvent may be any solvent typically used for catalytic hydrogenation, most typically a hydrocarbon or aqueous caustic. The catalyst used may be any catalyst typically used in homogeneous catalytic hydrogenations such as Pd(OAc)2 and Ru(PPh3)3Cl2, or in heterogeneous catalytic hydrogenations such as supported palladium, platinum, rhodium, ruthenium, nickel, sponge nickel and ZCuO.Cr2Os3 or a solid oxide such as celite, CuO, CrO3, CoO. SiOz, AI2O3, Ba(OH)2. MnO, At(iOPr)3, LnO3, ZrO and the zeolites. The reaction may be carried out at any temperature between 80oC and 300oC inclusive, preferably at a temperature between 180°C and 220°C inclusive. The hydrogen pressure may be any pressure below 50 bar, preferably between 5 and 35 bar inclusive. The catalyst loading may be between 0,01 and 20%, preferably between 0,05 and 5%. At the end of the step, the catalyst is removed or deactivated, and any solvent present is removed. This racemisation step may be earned out in conjunction with step (b), i.e the hydrogenation step, where appropriate. The next step, step (4) of the process of the invention is to hydrolyse the compound of the formula IV to the desired stereoisomer of the compound of the formula III. The reaction may be carried out in the presence of a base which may be a salt of a lower aliphatic alcohol such as sodium methoxide or sodium ethoxide, a metal hydroxide such as KOH, NaOH, or Mg(OH)2, or amine bases such as NH4OH. The reaction may be carried out in any solvent typically used in hydrolysis reactions, such as for example a lower aliphatic alcohol or water. Combinations of the solvents may also be used. . The reaction temperature may be any temperature below the boiling point of the chosen solvent or the reflux temperature of the mixture at the pressure at which the reaction is carried out. In the compound of the formula IV, where R1 is an isopropyl group, hydrolysis of this compound ((-)-menthyl ester) results in the production of (-)-menthoi. As a final step, the desired isomer of the compound of the formula III may be purified to the desired purity by, for example, distillation or crystallisation. Experimental Work The results of various experiments carried out in relation to the process of the invention are set out below. Example 1 m-Cresol (2,0 g) and o-phosphoric acid (4 mol e.g 8,5 g) were placed in a round bottom flask and heated to 85°C. Isopropano! (1,11 g) was added dropwise over 30 minutes. The reaction was then cooled to 25oC, and the organic phase extracted into toluene (20 ml). NMR analysis of the concentrated organic fraction indicated a 10% conversion of m-cresol to thymol. Example 2 Thymol (4,20 g), cyclohexane (50 ml) and 5% Pt/C (0,40 g) were placed in a 300 ml Parr autoclave reaction chamber. Once sealed, the chamber was flushed with N2 (g) before charging with H2 (g) (20 bar). The reaction was heated to 180oC and the reaction was allowed to proceed for 2 hours. The reaction was then allowed to cool to room temperature, the pressure inside the reactor was released and the reaction mixture filtered to give a mixture of menthol stereoisomers (100% conversion thymol, 80% selectivity to menthols). Example 3 In 2 ml vials, 10 or 100 mg of Pseudomonas cepacia lipase (Fluka). Pseudomonas fluorescens lipase (Fluka), penicillin acylase (Altus), subtilisin Carlsberg (Altus), Aspergillus oryzae protease and PeptiCLEC-TR slurry (Altus) were weighed. To this cyclohexane, hexane, pentane or heptane (962,72 μl) were added. Isomeric menthol (23 pi) and vinyl acetate (14,28 μl) were also added. These vials were incubated at 30'C or 37C for 2 or 48 hours. Enzyme was then removed from the mixture by centrifugation. Samples were analysed by gas chromatography (GC). An individual peak found or) each of the chromatograms was identified as that of (-)-menthyl acetate by comparison with the corresponding retention time of 2 standard sample of (-)-menthyl acetate. Example 4 Lyophilised Amano AK (a Pseudomonas fluorescens lipase enzyme) was obtained from Amano Pharmaceutical Co. (Japan). Concentrated liquid menthol was produced by the hydrogenation of thymol. The menthol contained four diastereomeric pairs of menthols, namely (±)-menthol (51%), (±)"isomenthol (14%), (±)-neomenthol (29%) and (±)-neoisomenthol (2%). A volume of 1ml was added to sealed batch reactors. Vinyl acetate (54 μl) was added at a 2:1 molar ratio to (-)-menthol. Heptane was added as solvent to a final reaction volume of 5 ml. These reactors were incubated in silicon oil baths at 50*C and stirred on a stirring hot plate. Batch times, unless otherwise stated were 24 hours. The reaction mixture was then centrifuged to separate the products from the enzyme. The supernatant was analysed by GC (%m/m analysis). Of the available (-)-menthol, 100% was converted to (-)-menthyl acetate, at an ee of 98%. The enzyme was recycled a total of 150 times by washing with heptane and adding fresh substrate (liquid menthol, vinyl acetate and heptane) after every recycle. An amount of menthyl acetate equivalent to 185 g of (-)-menthol was produced by this process. Example 5 Amano AK lipase enzyme was dissolved in phosphate buffer (5 mM, pH 7) before adding Celite 535 (in a ratio of 1:2). The mixture was frozen at -80°C for 5 minutes and then freeze-dried for 24 hours to produce immobilized enzyme. A substrate mixture containing 15,5% (v/v) of liquid menthol and 5,5% (v/v) vinyl acetate and 79% (v/v) of n-heptane was prepared. The mixture was pumped through 5 columns arranged in series packed with immobilised enzyme. The temperature of the reaction was maintained at 50°C. The end product was analysed for the formation of (-)-menthyl acetate by GC analysis- Of the available (-)-menthol, 100% was converted to (-)-menthyl acetate at an ee of 98%. In a 206 day period an amount of (-)"menthyl acetate equivalent to lOlg of menthol per gram of immobilized enzyme was produced by this immobilized enzyme system. Example 6 A solution of methanol (50,1 g), water (50,21 g) and NaOH (6,03 g, 0,15 mol) was placed in a reactor and heated to 60oC, (")-Menthyl acetate (30,53 g, 15 mol) was then added to the reactor and the reaction was vigorously stirred for 30 minutes. Qualitative GC analysis indicated a 53 area % menthyl acetate and a 47 area % menthol. Example 7 (")-Menthol (100 g) and Ni (1%) were placed in round bottom flask and heated at reflux for 2 hours. Once cooled to 25oC, the reaction mixture was sampled. Quantitative m/m analysis 17% thymol, 23% menthols and 54% menthones with a % (+)-menthol(±)-menthol of 22%. WE CLAIM: 1. A process of separating a single stereoisomer from a racemic mixture of eight stereoisomers of a compound of the formula ill t wherein R, represents an isopropanol group, an isopropyl group or an isopropylene group, Including the steps of; (1) contact the racemic mixture in a suitable organic solvent with an esterifying agent and a stereospecific enzyme which stereoseiectively esterifies the -OH group of the desired stereoisomer, for a time sufficient to convert a desired percentage of the desired stereoisomer to a compound of the formula IV without the reaction proceeding to the esterification of the other stereoisomers wherein R1 is as defined above and R2 is an alkyl or an aryi group, to give a first reaction product including the compound of the formula IV, the organic solvent, the unconverted stereoisomers of the compound of the formula 11!, excess esterifying agent and byproducts of the reaction; and (2) separating the compound of the formula IV from the first reaction product. 2. The process according to claim 1 wherein step (2) comprises the sub-steps of: (2)(a) separating the first reaction product from the enzyme; (2){b) removing from the first reaction product the organic solvent, the excess esterifying agent, and the by-products of the reaction to give a second reaction product including the compound of the formula IV and the unconverted stereoisomers of the compound of formula ill ; and (2)(c} separating the compound of the formula IV from the second reaction product leaving a third reaction product containing the unconverted stereoisomers of the compound of the formula HI, 3. The process according to claim 1 or claim 2 including the step, after step (2) of: (3) racemizjng the unconverted stereoisomers of the compound of the formula 111 in the third reaction product to give a fourth reaction product containing a mixture of all eight stereoisomers of the compound of the formula !il and recycling the fourth reaction product to step (1). 4. The process according to claim 3 including the step, after step (3) of: (4) hydrolysing the compound of the formula IV to give the desired stereoisomer of the compound of the formula 111, 5. The process according to claim 4 wherein when R1 is an isopropanol group or an isopropylene group, before or after step (4), the compound of the formula IV or the desired stereoisomer of the compound of the formula Hi is subjected to a reduction step to convert R1 to an isopropyl group. 6. The process according to any one of claims 1 to 5 including the following steps, prior to step (1) of: (a) reacting a compound of the formula I wherein R1 represents a methyl or a hydroxymethyl group, and R3 represents H, a base metal, a benzyl group or an ailyl group, with an alkylating agent in the presence of a catalyst \o give a compound of the formula i! wherein R1 is as defined in claim 1 and R2 and R3 are as defined above; and (b) hydrogenating the compound of the formula II in the presence of a catalyst to give a racemic mixture of the eight stereoisomers of the compound of the formula lil. 7. The process according to any one of claims 1 to 6 wherein in step (1) the solvent is selected from the group consisting of isooctane; n-heptane; decane; methyl cyclohexane; t-butyl methyl ether; xylene; kerosene; pentane; cydohexane; hexane; benzene; butano(; toluene; isopropanoi; ethyl lactate; and acetone. 8. The process according to claim 7 wherein in step (1) the solvent is n- heptane. 9. The process according to any one of claims 1 to 8 wherein in step (1) the esterifying agent is selected from the group consisting of vinyl acetate, butyl acetate, octanoic acid, isopropenyl acetate, vinyl butyrate, ethyl lactate and ethyl acetate. 10. The process according to claim 9 wherein in step (1) the esterify'tng agent is vinyl acetate. 11. The process according to any one of claims 1 to 10 wherein in step (1) the estenfying agent is used in a molar ratio to the single stereoisomer of the compound of the formula lil of 0.5:1 to 30:1 inclusive. 12. The process according to any one of claims 1 to 11 wherein in step (1) the stereospecific enzyme is used in an amount of from 1g/l to 60g/l inclusive of the mixture of the racemic mixture of the eight stereoisomers of the compound of the formula 111, the suitable organic solvent and the esterifying agent. 13. The process according to any one of claims 1 to 12 wherein step (1) is carried out at a temperature of from 20oC to 100oC inclusive and at atmospheric or higher pressure. 14. The process according to any one of claims 2 to 13 wherein after step (2)(a) the stereospecific enzyme is recycled to step (1). 15. The process according to any one of claims 2 to 13 wherein in step (1) the stereospecific enzyme is used in a continuous system wherein the stereo specific enzyme is retained within a reactor and wherein in step {2)(a) the first reaction product exits the reactor leaving behind the enzyme. 16. The process according to any one of claims 4 to 15 wherein in step (4) the hydrolysis is carried out in the presence of a base, in a suitable solvent, and at a temperature below the boiling point of the solvent or the reflux temperature of the mixture of reactants at the pressure at which the reaction is carried out. 17. The process according to any one of claims 1 to 16 wherein in the compound the formula III, R1 represents an isopropyl group. 18. The process according to claim 2 comprising (3) racemizing the unconverted stereoisomers of the compound of the formula III in the third reaction product to give a fourth reaction product containing a mixture of all eight stereoisomers of the compound of the formula III after step (2) and recycling the fourth reaction product to step (1)-

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