Enantioselective acylation/deacylation

Table 12. Enantioselectivities in the acylation and deacylation steps in the burst kinetics of the reaction of (Z)-Phe-PNP(52)...

The ratios of these slopes for L- and D-esters are shown in Table 12. The kL/kD values of the acylation step in the CTAB micelle are very close to those in Table 9, as they should be. It is interesting to note that the second deacylation step also occurs enantioselectively. Presumably it is due to the deacylation ocurring by the attack of a zinc ion-coordinated hydroxide ion which, in principle, should be enantioselective as in the hydroxyl group of the ligand. Alternatively, the enantioselectivity is also expected when the free hydroxide ion attack the coordinated carbonyl groups of the acyl-intermediate with the zinc ion. At any rate, the rates of both steps of acylation and deacylation for the L-esters are larger than those for the D-esters in the CTAB micelle. However, in the Triton X-100 micelle, the deacylation step for the D-esters become faster than for the L-esters. 

Using this approach, racemates of (27) were enantiomerically enriched using a lipase in organic solvent, followed by racemization of the unreacted enantiomer in buffer. Acylated derivatives (S)-(28) were obtained in yields >50% and >99% ee. Lipases with the opposite enantioselectivity produced (R)-28 in >99% ee. Subsequent chemical deacylation of (28) yielded enantiomerically enriched (27). 

The solvent present in biphasic reactions can still have an effect on the enzyme even though the enzyme functions primarily in an aqueous microenvironment. A particularly dramatic example is the lipase AH (lipase from Burkholderia cepac/fl)-catalysed desym-metrization of prochiral 1,4-dihydropyridine dicarboxylic esters, where either enantiomer can be accessed in high enantioselectivity by using either water-saturated cyclohexane or diisopropyl ether (DIPE) respectively (Scheme 1.60). The acyl group used in acylation and deacylation can also have a dramatic effect on enantioselectivity. " ... 

This process has many benefits in the context of green chemistry it involves two enzymatic steps, in a one-pot procedure, in water as solvent at ambient temperature. It has one shortcoming, however-penicillin acylase generally works well only with amines containing an aromatic moiety and poor enantioselectivities are often observed with simple aliphatic amines. Hence, for the easy-on/easy-off resolution of aliphatic amines a hybrid form was developed in which a hpase [Candida antarctica hpase B (CALB)] was used for the acylation step and peniciUin acylase for the deacylahon step [22]. The structure of the acyl donor was also optimized to combine a high enanhoselectivity in the first step with facile deacylation in the second step. It was found that pyridyl-3-acetic acid esters gave optimum results (see Scheme 6.8). 

Although many publications have covered the enantioselectivity of lipases in the deacylation step, their enantioselectivity in the acylation step (i.e., towards the acyl donor) has received much less attention. Generally, the selectivity of lipases towards racemic esters or acids is low to moderate [75-77]. Directed evolution and site-directed mutagenesis lead to a significant increase in the selectivity of the wild-type enzymes [78-80]. However, the enantiomeric ratios attained are still well below those typically obtained in kinetic resolutions of secondary alcohols. 

Enzymes are widely recognized as valuable tools for the synthesis of optically active compounds [22]. Thus, lipase-catalyzed acylation or deacylation is one of the most efficient methods for the preparation of optically active alcohols, acids, and esters. Because lipases retain activity and selectivity in non-conventional media such as organic liquids, their use as biocatalysts in enantioselective synthetic reactions has considerably increased. 

All three isomerizations discussed above seem to occur by analogous mechanistic pathways similar to the mechanism formulated for the Dakin-West reaction [82]. Deacylation of the starting material H by catalyst G affords, in a fast and reversible step (Scheme 13.47, step I), an acylpyridinium/enolate ion-pair I. From this ion pair, enantioselective C-acylation proceeds in the rate-determining and irreversible second step, furnishing the C-acylated product J (Scheme 13.47, step II). 

In order to broaden the scope we also examined [30] a combination of lipase-catalyzed acylation with penicillin acylase-catalyzed hydrolysis (deacylation). Good results (high enantioselectivity in the acylation and smooth deacylation) were obtained, with a broad range of both aliphatic amines and amines containing an aromatic moiety, using pyridylacetic acid ester as the acyl donor (Fig. 9.21). 

Enantioselective lipase-catalyzed transesterification involving deacylation of esters of racemic primary or secondary alcohols with primary alcohols, most frequently -butanol, serving as an acyl acceptor, is fairly common. Recent examples include esters of amino alcohols, isoserine, chlorohydrins, and various to-syloxybutanoate esters (eq 8). ... 

In the Novozym 435 catalysed ring-opening of a (chiral) substituted lacton, both acylation and deacylation can be enantioselective. For example, it is well known that CALB shows pronounced selectivity for / -secondary alcohols in the deacylation step. Since the forward and backward reaction exhibits by definition the same selectivity, esters comprising a substituent at the alcohol side are expected to show pronounced / -selectivity in the acylation step. " This is indeed observed for 7-MeHL, 8-MeOL and 12-MeDDL (Table 1). However, the selectivity for acyl donor in the case of PBL, 5-MeVL and 6-MeCL -lactones in which the ester bond is exclusively in a cisoid conformation- is low or for the S-enantiomer. We can speculate that lactones in a cisoid conformation must attain a different orientation in the active site in order to be activated. ... 

These findings led to elucidation of the mechanistic aspects of Upase (Novozym 435) catalysis enantioselection is operated by the deacylation step as shown in Fig. 3 [53], where only dimer formation is shown for simphcity. It is well accepted that at first the monomer (substrate) is activated by enzyme with formatimi of an (/ )-acyl-enzyme intermediate (enzyme-activated monomer, EM) [ acylation of lipase step (a) in Fig. 3]. Onto the activated carlxMiyl carbon of EM, the OH group of the D-lactate nucleophUically attacks to form an ester bond, liberating Upase enzyme and giving rise to D,D-dimer [ deacylation of Upase step (b) in Fig. 3]. 

The introduction of a substituent at the lactone ring inevitably generates a chiral center. As the action of lipases relies on a two-step mechanism with an acylation step and a deacylation step, involving a covalent acyl-enzyme intermediate vide supra), both the acylation and deacylation step can occur enantioselectively when using a (chiral) substituted lactone [41]. It is well known that lipases such as CALB show a pronounced selectivity for (R)-secondary alcohols in the deacylation step [42, 43]. Although less elaborately studied, the acylation step can also occur enantioselectively [44-50], and therefore the enzymatic ROP of substituted lactones may result in optically active polymers, since selectivity for one of the enantiomers can be expected. 

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