Enantioselective hydrolysis with Pseudomonas

PPL and Hpase from Pseudomonas sp. catalyze enantioselective hydrolysis of sulfinylalkanoates. For example, methyl sulfinylacetate (46) was resolved by Pseudomonas sp. Hpase in good yield and excellent selectivity (62). This procedure was suitable for the preparation of sulfinylalkanoates where the ester and sulfoxide groups are separated by one or two methylene units. Compounds with three methylene groups were not substrates for the Hpase (65). 

A very efficient and universal method has been developed for the production of optically pue L- and D-amino adds. The prindple is based on the enantioselective hydrolysis of D,L-amino add amides. The stable D,L-amino add amides are effidently prepared under mild reaction conditions starting from simple raw materials (Figure A8.2). Thus reaction of an aldehyde with hydrogen cyanide in ammonia (Strecker reaction) gives rise to the formation of the amino nitrile. The aminonitrile is converted in a high yield to the D,L-amino add amide under alkaline conditions in the presence of a catalytic amount of acetone. The resolution step is accomplished with permeabilised whole cells of Pseudomonas putida ATCC 12633. A nearly 100% stereoselectivity in hydrolysing only the L-amino add amide is combined with a very broad substrate spedfidty. 

Lipases have been used to effect the enantioselective esterification of cyanohydrins or the enantioselective hydrolysis of cyanohydrin esters. This works for aldehyde cyanohydrins. Selective (S)-cyanohydrin esterification is effected by the enzyme from Pseudomonas sp. [11], There is also an example of selective (R)-cyanohydrin esterification by Candida cylindracea lipase [12]. Effenberger has shown the feasibility of this approach in principle to produce a number of enantiopure cyanohydrins derived from aldehydes. In situ derivatization with racemization as shown in Fig. 7 is possible, resulting in theoretically 100% yield of the desired enantiomer [13]. Ketone cyanohydrins, which are tertiary alcohols, do not easily undergo this reaction. 

Such a process, which runs in water and is based on a palladium-catalyzed racemization of O-acylated allyUc alcohols in combination with a hydrolase-catalyzed enantioselective hydrolysis of the ester, was developed in an early pioneering work by Allen and Williams [11]. In this type of DKR process, the resulting allylic alcohols of type, for example, (S)-6, were obtained with satisfactory to high enan-tioselectivities (Scheme 19.3). For example, under these conditions, the hydrolysis of racemic ester rac-5 in the presence of a lipase from Pseudomonas Jluorescms gave... 

Enantioselective hydrolysis of nitriles into amides or acids has primarily been catalyzed by various Rhodococus and Pseudomonas species (Fig. 10.33). Prochiral compounds were also hydrolyzed to give the corresponding acid in high yield and ee (Fig. 10.33(d), (e) and (g)). - In the reaction of substituted malononitrile with Rhodococcus rhodochrous, the first hydrolysis step leading to diamide proceeded without enantiodiscrimination, but further hydrolysis of the diamide proceeded with high enantioselectivity, affording the (Placid in 92% yield and 96% ee (Fig. 10.33(g)). 

For the synthesis of p-lactam antibiotics, the presence of asymmetrical carbon at the 3 and 4 positions is critical to prepare optically active -lactams [197]. Nagai et al. [198] developed enzymatic synthesis of optically active p-lactams by lipase-catalyzed kinetic resolution using the enantioselective hydrolysis of iV-acyloxymethyl p-lactams 108 in an organic solvent (isopropyl ether saturated with water) and the transesterification of N-hydroxymethyl P-lactam 109 in organic solvent (metiiylene chloride) in tiie presence of vinyl acetate as acyl donor (Fig. 37). The reaction yield of 35-50% and e.e. s of 93 to more than 99% were obtained depending on the specific substrate used in the reaction mixture, Lipase B from Pseudomonas fragi and lipase PS-30 from Pseudomonas sp. were used in the reaction mixture. 

Two more examples ia Table 5 iaclude the hydrolysis of esters of trans-alcohols that proceed with high efficiency practically regardless of the nature of the substituents (72) and resolution of P-hydroxynitriles with Upase from Pseudomonas sp. In the latter case the enantioselectivity of the hydrolysis was improved by iatroduciag sulfur iato the acyl moiety (73). 

Pseudomonas aeruginosa lipase-catalyzed hydrolysis of racemic ester 23 proceeds with very low enantioselectivity E = 1.1). Sequential use of error-prone PCR, saturation mutagenesis at chosen spots and DNA shuffling resulted in the formation of a mutant whose enantioselectivity was over 50. 

Early reports on the effects of the choice of solvent on enzymatic enantioselectivity showed that substantial changes may be observed. For the transesterification reaction of sec-phenethyl alcohol with vinyl butyrate catalyzed by subtilisin Carlsberg, a 20-fold increase in the E-value was reported when the medium was changed from acetonitrile to dioxane [59]. Similar changes were recorded for the prochiral selectivity of Pseudomonas sp. lipase in the hydrolysis of 2-substituted... 

Diols such as the optically active 1,1 -binaphthyl-2-2 -diol (BINOL) have been used as versatile templates and chiral auxiliaries in catalysts employed successfully in asymmetric synthesis. The application of enzymes in the enantioselective access to axially dissymmetric compounds was first reported by Fujimoto and coworkers.83 In aqueous media, the asymmetric hydrolysis of the racemic binaphthyl dibutyrate (the ester) using whole cells from bacteria species afforded the (A)-diol with 96%ee and the unreacted substrate (A)-ester with 94% ee at 50 % conversion. Recently, in non-aqueous media, lipases from Pseudomonas cepacia and Ps. fluorescens have been employed in the enantioselective resolution and desymmetrization of racemic 6,6 -disubstituted BINOL derivatives using vinyl acetate.84 The monoacetate (K)-73 (product) was obtained in 32-44 % chemical yields and 78-96% ee depending on the derivatives used. The unreacted BINOL (S)-72 was obtained in 30-52 % chemical yield and 55-80% ee. 

DSM developed a slightly different approach towards enantiopure amino acids. Instead of performing the Strecker synthesis with a complete hydrolysis of the nitrile to the acid it is stopped at the amide stage. Then a stereoselective amino acid amidase from Pseudomonas putida is employed for the enantioselective second hydrolysis step [83], yielding enantiopure amino acids [34, 77, 78]. Although the reaction is a kinetic resolution and thus the yields are never higher than 50% this approach is overall more efficient. No acylation step is necessary and the atom efficiency is thus much higher. A drawback is that the racemisation has to be performed via the Schiff s base of the D-amide (Scheme 6.23). 

Another remarkable example of a solvent-induced ehange in enantioselectivity is the partial hydrolysis of achiral 4-aryl-substituted l,4-dihydro-2,6-dimethyl-3,5-pyridine diearboxylie diesters, catalysed by a lipase from Pseudomonas sp., leading either to the (P)- or (5)-eonfigured monoesters. Whereas in water-saturated di-/-propyl ether, the S)-monoesters are obtained with ee values up to 99%, the (P)-monoesters are formed in water-saturated eyclohexane with ee values of 88-9 l%i [316]. 

The lipase from Pseudomonas aeruginosa (PAL) catalyzes the hydrolysis of 2-me-thyldecanoic acid p-nitrophenyl ester with only 2% ee in favor of the (S)-acid. Reetz and Jaeger used four rounds of error-prone PCR and screening on enantiomerically pure R and S substrates to generate a more enantioselective variant that produced the desired (S)-acid with 81% eell57l Additional cycles of error-prone PCR in combination with saturation mutagenesis further improved the enantioselectivity of this enzyme, which hydrolyzes the 2-methyldecanoic acid p-nitrophenyl ester with 91 % ee (E = 25.8) in favor of the (S)-acid 1Z. ... 

Prochiral diketones or racemic ketones, like enol esters, are also amenable to a hydrolase-catalyzed asymmetric transformation. The enol acetates and ketones 63 and 64, respectively, may be obtained by Pseudomonas cepacia lipase-catalyzed and Candida cylindracea lipase-catalyzed hydrolysis of the corresponding racemic enol esters or prochiral bis enol ester, respectively, with high enantioselectivity and yield. 

A series of alkyl, alkoxy or acylamino 1,3-proanediol derivatives substituted in 2-position have been subjected to lipase-catalyzed acylation, and the monoacetates (1-12, 19, 20, 23-38, 40-42) were obtained with moderate to high enantiomeric excess (Table 11.1-17). For the monoacetates 1-12, reactions with and in ethyl acetate are usually slower than those with and in vinyl acetate. As in the hydrolysis of the corresponding diacetates, much higher selectivities were recorded with the yet unidentified carboxyl esterase from crude pig pancreas lipase. An excellent lipase for the enantioselective acylation of 3-benzyloxy-l,3-propane diol is Pseudomonas fluor-escens lipase, which gives high selectivity with vinyl acetate, isopropenyl acetate and ethyl acetate. By carrying the acylation further, to a certain extent to the diacetate, the enantiomerically pure monoacetate should be obtainable. 

Most remarkably, a series of primary alcohols whose chiral center bears only alkyl or alkenyl groups (23-30) has been obtained with high enantioselectivity through Pseudomonas fluorescens lipase-catalyzed acylation with vinyl acetate in dichloromethane. For the attainment of chiral primary alcohols, lipase-catalyzed acylation seems to be more efficient in terms of selectivity and yield than lipase-catalyzed hydrolysis of the corresponding esters. A comparison ofTables 11.1-19 and... 

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