Esterification, kinetic resolution, lipases

Orthoformates have been used in the lipase-catalyzed esterification aimed at the kinetic resolution of racemic acids such as flurbiprofen, a nonsteroidal anti-inflammatory drug (Figure 6.18). Orthoformates trap the water as it is formed through hydrolysis, and therefore prevent the reverse reaction, and, at the same time, provide the alcohol for the esteriflcation [65]. 

The ability of enzymes to achieve the selective esterification of one enantiomer of an alcohol over the other has been exploited by coupling this process with the in situ metal-catalysed racemisation of the unreactive enantiomer. Marr and co-workers have used the rhodium and iridium NHC complexes 44 and 45 to racemise the unreacted enantiomer of substrate 7 [17]. In combination with a lipase enzyme (Novozyme 435), excellent enantioselectivities were obtained in the acetylation of alcohol 7 to give the ester product 43 (Scheme 11.11). A related dynamic kinetic resolution has been reported by Corberdn and Peris [18]. hi their chemistry, the aldehyde 46 is readily racemised and the iridium NHC catalyst 35 catalyses the reversible reduction of aldehyde 46 to give an alcohol which is acylated by an enzyme to give the ester 47 in reasonable enantiomeric excess. 

Catalytic sol-gel lipase immobilizates were rapidly commercialized (by Fluka) after their invention in 1995 because of their remarkably stable activity in esterification reactions (and also in the kinetic resolution of chiral alcohols and amines) along with unique stability (residual activity of 70% even after 20 reaction cycles is common). The original procedure for the encapsulation produced by the fluoride-catalysed hydrolysis of mixtures of RSi(OCH3)3 and Si(OCH3)4 has been improved... 

The one-pot dynamic kinetic resolution (DKR) of ( )-l-phenylethanol lipase esterification in the presence of zeolite beta followed by saponification leads to (R)-l phenylethanol in 70 % isolated yield at a multi-gram scale. The DKR consists of two parallel reactions kinetic resolution by transesterification with an immobilized biocatalyst (lipase B from Candida antarctica) and in situ racemization over a zeolite beta (Si/Al = 150). With vinyl octanoate as the acyl donor, the desired ester of (R)-l-phenylethanol was obtained with a yield of 80 % and an ee of 98 %. The chiral secondary alcohol can be regenerated from the ester without loss of optical purity. The advantages of this method are that it uses a single liquid phase and both catalysts are solids which can be easily removed by filtration. This makes the method suitable for scale-up. The examples given here describe the multi-gram synthesis of (R)-l-phenylethyl octanoate and the hydrolysis of the ester to obtain pure (R)-l-phenylethanol. 

Lipase-catalysed esterification instead of ester hydrolysis is also being used, a.o. by Glaxo Wellcome for a kinetic resolution that yields (IS, 2. -tranx-2-methoxycyelo-... 

Enzymes such as pig liver esterase have been successfully applied in enantioselective hydrolysis of allenyl esters on a scale of 2 mmoles131. This provides the enantiomerically enriched allene-carboxylic acid as well as the ester of opposite configuration, by what is in fact a catalytic kinetic resolution (6-90% oy). Conversely, partial enantioselective esterification of /1-hydroxy-allenes (3-72% oy) employing lipases has been reported132,133. 

Dynamic kinetic resolutions of secondary alcohols and amines have been achieved by the combination of biocatalysts with metal catalysts.12 For example, a metal catalyst was used to racemize the substrate, phenylethanol, and a lipase was used for the enantioselective esterification as shown in Figure 12. The yield was improved from 50% in kinetic resolution without racemization of the substrate to 100% with metal catalyzed racemization. 

There are many reports of enzymatic catalysis in scC02 performing hydrolysis, oxidations, esterifications, and franr-esterification reactions. For example, the enzymatic kinetic resolution of 1-phenylethanol with vinyl acetate in scC02 using lipase from Candida antarctica B produces (R)-l-phenyethylacetate in >99% ee (i.e., enantiomeric excess, a measure of how much of one enantiomer is present as compared to the other), as shown in Figure 12.20. 

Lipases have been extensively used for the kinetic resolution of racemic alcohols or carboxylic acids in organic solvents. Chiral alcohols are usually reacted with achiral activated esters (such as vinyl, isopropenyh and trichloroethyl esters) for shifting the equilibrium to the desired products and avoiding problems of reversibility. For the same reasons, chiral acids are often resolved by using acidolysis of esters. In both cases, the overall stereoselectivity is affected by the thermodynamic activity of water of water favors hydrolytic reactions leading to a decrease in the optical purity of the desired ester. Direct esterifications are therefore difficult to apply since water formed during the reaction may increase the o of the system, favors reversibiUty, and diminishes the overall stereoselectivity. 

Figure 7.2 The structure of the faster reacting enantiomer in lipase-catalyzed esterification in kinetic resolution of racemic secondary alcohols or hydrolysis of the corresponding esters. Small and large refer to the relative size of the groups and not to the R/S notation.

It should be evident that the maximum yield of a particular enantiomer normally available from a racemic mixture is 50%. However, in some enzymic catalysed kinetic resolutions it is possible to obtain >50% yield of one enantiomer from a racemate. For this to occur, it is necessary to have the desired chemical reaction, e.g. enzyme-catalysed stereoselective esterification, occurring at the same time as the enantiomers of the racemic starting compound are interconverting under equilibrium conditions. A successful example of this technique is provided by ben-zaldehyde cyanhydrin (2-hydroxy-2-phenylacetonitrile), whose R and S enantiomers, 49 and 50, respectively, equilibrate in the presence of a basic anion-exchange resin (Scheme 3.5). In the presence of lipase, (S)-ben-zaldehyde cyanhydrin acetate 51 was formed in 95% yield and in 84% enantiomeric excess (see Inagaki et al.u and Ward15). 

Other possibilities to prepare chiral cyanohydrins are the enzyme catalysed kinetic resolution of racemic cyanohydrins or cyanohydrin esters [107 and references therein], the stereospecific enzymatic esterification with vinyl acetate [108-111] (Scheme 2) and transesterification reactions with long chain alcohols [107,112]. Many reports describe the use of fipases in this area. Although the action of whole microorganisms in cyanohydrin resolution has been described [110-116],better results can be obtained by the use of isolated enzymes. Lipases from Pseudomonas sp. [107,117-119], Bacillus coagulans [110, 111], Candida cylindracea [112,119,120] as well as lipase AY [120], Lipase PS [120] and the mammalian porcine pancreatic lipase [112, 120] are known to catalyse such resolution reactions. 

Scheme 2. Kinetic resolution of racemic cyanohydrins by lipase catalysed hydrolysis and by lipase catalysed irreversible esterification with vinyl acetate...

Lipase-catalyzed esterifications of racemic carboxylic acids in SCFs have been studied by several groups. The target in all of these studies was the preparation of optically pure anti-inflammatory drugs ibuprofen and naproxen. Rantakyla and Aaltonen reported the kinetic resolution of racemic ibuprofen by esterification catalyzed by immobilized lipase from Mucor miehei [Eq. (1)] (8,74,75) ... 

Kinetic resolution of chiral alcohols by selective esterification has also been performed in SCCO2 by several groups. A glass-immobilized lipase from Candida cylindracea was found to catalyze the stereoselective esterification of only the S isomer from racemic citronellol (Eq. (3)] in a continuous-flow reactor (78) ... 

Because free-enzyme porcine pancreatic lipase had much lower activity in SCCO2 than in organic solvents, the authors decided instead to test immobilized lipases. Macroporous resin-supported lipase from M. miehei had fair activity but poor enantioselectivity. Porcine pancreatic lipase immobilized on supports had the greatest activity if the support was highly hydrophilic Sephadex G-25 and Bio-gel P6 were selected. Enantioselectivity was 83% for the (5)-gycidyl butyrate at 25-30% conversion, comparable to results in organic solvents. Several chiral alcohols were studied by Cernia et al. (80) as substrates for kinetic resolution by esterification catalyzed by silica-supported lipase from Pseudomonas sp. [Eq. (5)] ... 

On the other hand if the acid is the interesting part then the same two types of reaction can be carried out to produce enantiomerically enriched esters or acids as required. Both these reactions are equilibria. Both are also kinetic resolutions (chapter 28) and it is not usually possible to get both products in high ee. The key statistic is the rate ratio ( E ) of hydrolysis or esterification of the two enantiomers. Since the molecules we shall be using are not natural substrates of lipases, the E value will vary considerably but, as you know from chapter 28, it does not need to be very large for acceptable results. This E is often called ks/kR in chemical kinetic resolution and called the s value in chapter 28. 

Resolution by transesterification. Using vinylic acetates to esterify allyl alcohols, propargyl alcohols, 2-phenylthiocycloalkanols, a-hydroxy esters," methyl 5-hydroxy-2-hexenoates, and 2-substituted 1,3-propanediols, the enantioselective esterification provides a means of separation of optical isomers. Vinyl carbonates are also resolved by lipase-mediated enantioselective conversion to benzyl carbonates. Other esters that have also been used in the kinetic resolution include 2,2,2-tri-fluoroethyl propionate. There is a report on a double enantioselective transesterification" of racemic trifluoroethyl esters and cyclic meso-diols by lipase catalysis. 

Pentanediol [(R,/ )-28] has been used for the formation of chiral acetals and as a precursor for chiral alkenes (Sections D.1.5.1. and 1.6.1.5.). The original procedure for the resolution of the phenylboronic acid derivative with brucine39 was impractical, but recently a kinetic resolution by lipase-catalyzed esterification and hydrolysis has been developed40. In addition, a good method for catalytic reduction of 1,3-diketones with Raney nickel modified with sodium bromide and tartaric acid (for a procedure, see Section D.2,3.1.) allowed the production for commercial purposes41. Similarly, sterically more hindered and less water-soluble 2,6-dimethyl-3,5-heptanediol (29) has been introduced for the same purpose. It is obtained in the same way from the diketone and separated from the meio-compound by simple recrystallization42. 

Of the six main classes of enzymes, hydrolases, oxidoreductases and transferases have been the three most useful in kinetic resolution. Among the hydrolases, lipases are extensively used. The molecular machinery of lipases consists of a catalytic triad of the amino acids serine, histidine, and aspartic (or glutamic) acid. The enzyme first transfers the acyl group of an ester to the hydroxyl group of the serine residue to form the acylated enzyme. The acyl group is subsequently transferred to an external nucleophile with the return of the enzyme to its pre-acylated state to start the process again. A variety of nucleophiles can participate in this process water results in hydrolysis, an amine results in amidation, an alcohol results in esterification or transesterification, and hydrogen peroxide results in the formation of perac-id. Another reason which favored the relatively wide applicability of lipases in enzymatic... 

In the context of continuous-flow operations using SCCO2/ILS, it has to be mentioned that this concept has also been successfully applied to biocatalyzed reactions. Among others, Reetz et al. were able to demonstrate that kinetic resolution of alcohols is possible by lipase-catalyzed alcohol esterification in ILs and subsequent continuous extraction of the ester derivatives with SCCO2 [29]. For further information, see Chapter 8. 

Lipases and esterases are often used for the kinetic resolution of racemates either by hydrolysis, esterification or transesterification of suitable precursors. (Scheme 8-4) illustrates the principle for the resolution of secondary alcohols by esterification with vinyl acetate... 

Dynamic Kinetic Resolution. Another typical acid-catalysed reaction is the racemisation of chiral alcohols, due to inversion at the chiral carbon. This can actually be made use of in the formation of enantiopure compounds, by dynamic kinetic resolution using an enzyme, such as a lipase, that catalyses enantioseleetive esterification in an organic medium. By coupling zeolite Beta-catalysed intereonversion of benzylic alcohol enantiomers with enzyme-catalysed esterifieation of only one of the enantiomeric alcohols, almost complete eon version to enantiopure ester ean be achieved. ... 

An elegant combination of monomers with the components of a dynamic kinetic resolution (DKR) permitted the conversion of a racemic diol into a polymer consisting of enantioenriched units that could be recovered by polymer hydrolysis [28]. Diol 5 and achiral diester 6 were combined with a well-known system of lipase and ruthenium catalyst (see Chapters 4 and 5 for more on this). The esterification of the free hydroxyl groups is very selective (for the R) configuration) but as the polymerization proceeds, the (S) stereocentres are racemized. Upon 92% conversion of the hydroxy groups and hydrolysis of the polymer, an enantioenriched sample of the diol was obtained that contained essentially none of the (S,S)-isomer. 

The Mitsunobu esterification was the first inversion of configuration applied together with the lipase-catalyzed acylation of secondary alcohols (38). Another possibility is shown in Figure 23. The CAL-B-catalyzed kinetic resolution of ethyl-3-hydroxybutanoate by acylation with vinyl acetate has been discussed already in Figure 4 (9). Normal kinetic resolution under the solvent-free conditions by... 

Lipases are used to catalyze hydrolytic, esterification and transesterification reactions. These reactions alter the physical properties of fats and oils and thereby produce a wide range of products (Mukherjee, 1990). Lipases have also been used for the kinetic resolution of Isomers of alcohols or fatty acids (Hills, et 1990). The lipases... 

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