Amino acid ethyl esters, resolution

Finally, as an old example of kinetic resolution of racemic mixtures, mention must be made on the report of Kise and Tomiuchi on the significant effect of acetonitrile on the enantioselectivity of different proteases toward the kinetic resolution of aromatic amino acid ethyl esters (5-8). For instance, (l)-DOPA (8) was obtained with 99% ee in the presence of 90% v/v acetonitrile [9]. 

SaUcylaldehydes have been employed to assist in the racemization ofa-amino acid ethyl esters. Condensation of the amino acid ester with the salicylaldehyde derivative led to an increase in the acidity of the a-hydrogen atom (at the stereogenic centre) leading to rapid racemization. The DKR procedure employed an endoproteinase alcalase enzyme for the resolution step and various aldehydes for the in situ racemisation step. Hydrolysis of the imine liberated the amino acid ester and the aldehyde. The a-amino acids thus obtained in high yield (up to 94%) exhibited high enantiopurity (up to 98%) (Schemes 4.20 and 4.21) [56]. 

In addition to the mobile phase composition, the effect of other parameters such as temperature, flow rate, pH, and structure of the analytes were also studied, but only a few reports were available in the literature. In 1995, Lin and Maddox [66] studied the effect of temperature on the chiral resolution of amino acids and esters. The temperature was varied from 5°C to 25°C and it was reported that the resolution improved at low temperature. Hyun et al. [48-50,67] carried out the effect of temperature on the chiral resolution of amino alcohols, amines, fluoroquinolones, and other drugs. Again, lowering of temperature resulted in better resolution. The effect of temperature on the chiral resolution of phenylalanine, phenylglycine, and 2-hydroxy-2-(4-hydroxy-phenyl)-ethyl amine is shown in Table 5 [50], which indicates an increase in retention factors at lower temperature, but the best separation occurred at 20°C. These experiments indicated the exothermic nature of chiral resolution on CCE-based CSPs. Lin and Maddox [66] also studied the effect of flow rate on the chiral resolution of... 

One Sanofi synthesis of enantiomerically pure (-i-)-clopidogrel (2) utilized optically pure (R)-(2-chloro-phenyl)-hydroxy-acetic acid (20), a mandelic acid derivative, available from a chiral pool. After formation of methyl ester 21, tosylation of (/ )-21 using toluene sulfonyl chloride led to a-tolenesulfonate ester 22. Subsequently, the Sn2 displacement of 22 with thieno[3,2-c]pyridine (8) then constructed (-i-)-clopidogrel (2). Another Sanofi synthesis of enantiomerically pure (-i-)-clopidogrel (2) took advantage of resolution of racemic a-amino acid 23 to access (S)-23. The methyl ester 24 was prepared by treatment of (S)-23 with thionyl chloride and methanol. Subsequent Sn2 displacement of (2-thienyl)-ethyl para-toluene-sulfonate (25) assembled amine 26. 

Initial scale up of the enzymatic resolution for production of kilogram quantities of (R)-2-amino-2-ethylhexanoic acid was performed in a batch process. The oil of ethyl 2-amino-2-ethyl-hexanoate was suspended in an equal volume of water containing the enzyme. The enantioselective hydrolysis of the ester proceeded at room temperature with titration of the produced acid by NaOH through a pH stat (Figure 6.4). 

The selective resolution enhancement in derivative spectroscopy is pushed even further in the fourth derivative mode. As in the case of second derivative spectroscopy, the amplitude and the position of the derivative spectral bands of the aromatic amino acids are related to the polarity of the medium. We have undertaken a systematic investigation of these spectral features of the N-acetyl O-ethyl esters of tyrosine and tryptophan in various solvents of different polarity (from cyclohexane to water). Astonishingly, a simple relationship between the spectral parameters of the fourth derivatives and the dielectric constant was found [11]. As shown in Figure 5, for tyrosine it is the position of >.max, and for tryptophan it is the derivative amplitude which depends linearly on the dielectric constant er. Since in addition the fourth derivative spectra of these model compounds do not depend significantly on pressure (at least up to 500 MPa), these spectral features may be used as an intrinsic probe to sense the dielectric constant in the vicinity of tyrosine and tryptophan. 

Kanerva et al. resolved ethyl esters of ten alicyclic (i-aminocarboxylic acids by lipase catalysis in organic solvents. The resolutions were based on acylation of the amino group at the / -stereogenic centre with various 2,2,2-trifluoroethyl esters. From the cis and trans racemic esters 42, all four enantiomers of 2-ACPC were prepared. The absolute configurations of 43 and 44 were proved by transformation to the known 2-ACPC enantiomers by hydrolysis and subsequent desalting with an anion-exchange resin [85]. 

Enzymes other than CAL B have also been reported to operate under the biphasic conditions. CAL A and CAL B or a lipase from Mucor miehei were tested for the kinetic resolution of glycidol using vinyl acetate or vinyl butyrate. The enzymes were used either suspended in the free ILs or immobilized, when the reactions were carried out in [EMIM][NTf2] [71]. CALAwas inactive, butthe other two enzymes showed activity, albeit at 10-20% of that in the absence of COj whether they were free or immobilized. In general, the supported enzymes showed superior performance [71]. Chymotripsin, a specific protease for aromatic amino acids, was found to catalyze the hydrolysis or transesterification of the ethyl ester of N-acetyl-phenylalanine with propanol in scC02-[RMIM][PF5] (R = butyl or octyl) with or without added water [Eq. (15)]. 

An important question in molecular imprinting has been addressed using covalent binding by two boronic acids to what extent can imprinted polymers also bind substances other than the template. For example, are racemates of other substances resolvable In the first experiments on glyceric acid esters 5 with a certain ester as template, imprinted polymers were shown to resolve a whole series of racemates even when the alcohol group in the racemate is varied (methyl, ethyl, benzyl, or 4-nitrophenyl) [39]. Aromatic amino acids were shown to behave similarly. Here, the aromatic group in the racemates can vary. A racemate resolution is possible provided that the rest of the structure remains the same [40]. 

There are other published resolutions of 4-hydroxyphenylglycine, one of which involves the enzyme-catalysed hydrolysis of the ethyl ester (Scheme 6.9). Since the substrate is fully blocked at the amino and the carboxylate functions, it is scarcely soluble in water and must be dissolved in an organic solvent. The specific hydrolysis of the ester catalysed by an enzyme in an aqueous phase in contact with the organic solvent will yield a water-soluble carboxylic acid which transfers to the aqueous phase containing the enzyme. The conditions of the reaction therefore effect both hydrolysis of the ester and facile separation of the product. In fact, the enzyme is immobilized on a hydrophilic membrane at the interface between the two immiscible phases. The membrane reactor (Figure 6.2) comprises a large bundle of hollow fibres, each having an external diameter of about... 

The resolution of amino acids was reported employing the ionic Hquid 1-ethyl-3-methylimidazolium acetate. Esters of various DL-amino acids were used and DL-phenylalanine methyl ester exhibited the highest ee (>98%). An interestingly high ee (98.2%) was also found for the resolution in deuterium oxide (D2O) as opposed to ordinary water and it was suggested that this was caused by protein stabihzation by the DjO [20]. 

Zhao, H. and Malahorta, S. V., Enzymatic resolution of amino acid esters using ionic liquid N-ethyl pyridinium trifiuoroacetate. Biotech. Lett., 24 1257-1260, 2002. 

The first DKR of a-amino acids was achieved using their amides as the substrate, Novozym 435 as the resolution enzyme, Pd/A10(0H) as the racemization catalyst, and ethyl acetate as the acyl donor [46]. The DKR of phenylalanine amide was successful at 100 °C (Scheme 5.37). In contrast to this, the DKR of phenylglycine amide and simple derivatives was achieved at lower temperature (60 °C) [60]. On the other hand, the use of N-cbz-glydne and N-cbz-gly-glycine esters as the acyl donors provided enantioenriched dipeptides and tripeptides (Chart 5.33). 

Further studies on L-Phe-An MIP plates with the enantiomers of various compounds, such as anilide, dansyl, methyl and ethyl esters, and amide derivatives of amino acids, showed that polymer interacts more weakly with enantiomers of these compounds and, in the case of dansyl derivatives, no chiral resolution was observed, probably due to the dansyl group that blocks the ammine group of phenylalanine. 

Resolution of a,a-disubstituted a-amino acids (but also of a,a-disubstituted a-hydroxy acids [51]) can also be performed on their esters. We have used pig liver esterase (PLE) for resolution of a variety of a,a-disubstituted esters. Although all substrates tested are hydrolyzed, only for a-substituted phenylglycine esters (17, X = NH2) and a-substituted mandelic esters (17, X = OH) reasonable enanlioselectivities are observed for hydrolysis of the (S) enantiomer (E = 2-114) [44]. For these types of amino acids and hydroxy acids the PLE resolution forms a valuable extension of the M. neoaurum resolution technology, since the corresponding racemic amides are hydrolyzed sluggishly (Scheme 8). The PLE resolution of a-allylphenylglycine ethyl ester has been applied in the synthesis of D-a-phenylproline (vide infra). 

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