Deracemization of Amines

Scheme 2.31 Deracemization of amines via combined use of an enantioselective amine oxidase and ammonia borane.

Oxidoreductases are, after lipases, the second most-used kinds of biocatalysts in organic synthesis. Two main processes have been reported using this type of enzymes-bioreduction of carbonyl groups [39] and biohydroxylation of non-activated substrates [40]. However, in recent few years other processes such as deracemization of amines or alcohols [41] and enzymatic Baeyer-Villiger reactions of ketones and aldehydes [42] are being used with great utility in asymmetric synthesis. 

The group of Turner has reported the deracemization of amines [79]. The wild type of Type II monoamine oxidase from Aspergillus niger possesses very low but measurable activity toward the oxidation of L-a-methylbenzylamine. The oxidation of the D enantiomer is even slower. In vitro evolution led to the identification of a mutant with enhanced enantioselectivity, showing high E values (>100) for a variety of primary and secondary amines. An example is shown in Scheme 5.39. 

TABLE 2.7 One-Pot Simultaneous Deracemization of Amines Combining (S)- and (/ )-Selective w-TAs ... 

Subsequently Turner and coworkers were able to show that the Asn336Ser variant possessed broad substrate specificity, with the ability to oxidize a wide range of chiral amines of interest [19]. They also discovered a second mutation, Ile246Met, which conferred enhanced activity toward chiral secondary amines as exemplified by the deracemization of racemic 1-methyltetrahydroisoquinoline (MTQ) (9) (Figure 5.9)[20j. 

Upon mutagenesis of the monoamine oxidase from Aspergillus niger (MAO-N) within several rounds of directed evolution [65], variant biocatalysts were identified with largely expanded substrate acceptance, enabling also the deracemization of tertiary amines incorporating straight-chain and cyclic structural motifs [66]. 

Figure 11.1 Enzymatic deracemization of racemic amines via a two-step, one-pot process utilizing an enantioselective amine oxidase in combination with ammonia-borane.

In order to extend the approach to include deracemization of chiral secondary amines, this group carried out directed evolution on the monoamine oxidase (MAO) enzyme MAO-N (Scheme 2.32). A new variant was identified with improved catalytic properties towards a cyclic secondary amine 64, the substrate used in the evolution experiments. This new variant had a single point mutation, lle246Met, and was found to have improved catalytic properties towards a number of other cyclic secondary amines. The new variant was used in the deracemization of rac-64 yielding (R)-64 in high yield and enantiomeric excess [34]. 

An elegant four-enzyme cascade process was described by Nakajima et al. [28] for the deracemization of an a-amino acid (Scheme 6.13). It involved amine oxidase-catalyzed, (i )-selective oxidation of the amino acid to afford the ammonium salt of the a-keto acid and the unreacted (S)-enantiomer of the substrate. The keto acid then undergoes reductive amination, catalyzed by leucine dehydrogenase, to afford the (S)-amino acid. NADH cofactor regeneration is achieved with formate/FDH. The overall process affords the (S)-enantiomer in 95% yield and 99% e.e. from racemic starting material, formate and molecular oxygen, and the help of three enzymes in concert. A fourth enzyme, catalase, is added to decompose the hydrogen peroxide formed in the first step which otherwise would have a detrimental effect on the enzymes. 

An important breakthrough was made very recently in this area. A chemoenzymatic method developed by Turner has allowed the cyclic deracemization of tertiary amines [80]. Enantiopure tertiary amines cannot be obtained via DKR. One of the variants obtained by directed evolution of the monoamine oxidase from Aspergillus niger showed high activity and enantioselectivity toward cyclic tertiary amines (Scheme 5.40). 

The deracemization of a number of pharmaceutically valuable building blocks has been carried out by biocatalytic processes. This includes epoxides, alcohols, amines and acids. DKR involves the combination of an enantioselective transformation with an in situ racemisation process such that, in principle, both enantiomers of the starting material can be converted to the product in high yield and ee. The racemization step can be catalysed either enzymatically by racemases, or non-enzymatically by transition metals. 

Carr, R., Alexeeva, M., Dawson, M. J., Gotor-Fernandez, V., Humphrey, C. E., and Turner, N. J. 2005. Directed evolution of oxidase for the preparative deracemization of cyclic secondary amines. Chem. Bio. Chem.,6, 637-639. 

The method is of general applicability in the deracemization of secondary alcohols and amines and consists of a Upase-catalyzed irreversible acylation and in situ racemization of the non-reacted enantiomer catalyzed by a ruthenium catalyst. 

The coupling of these two enzymatic systems could find many more applications due to the avaUabihty of amino acid dehydrogenases of broader specificity [31]. A series of amino acid dehydrogenases with D-specificity for the preparation of D-amino acids has been applied to the reductive amination of a-keto acids [32]. However, the deracemization of rac-amino acids exploiting this type of enzyme requires an amino acid oxidase with L-specificily, which is a rare enzymatic activity. As an alternative the a-oxo acid, usually available through difficult synthetic procedures, can be used directly. 

Figure 14.34 Deracemization of racemic amines by repeated cycles of enzyme catalyzed enantioselective oxidation followed by nonselective chemical reduction.

Enzymatic oxidations of carbon-nitrogen bonds are as diverse as the substances containing this structural element. Mainly amine and amino acid oxidases are reported for the oxidation of C-N bonds. The steroespecificity of amine-oxidizing enzymes can be exploited to perform resolutions and even deracemizations or stereoinversions (Fig. 16.7-1 A). Analogous to the oxidation of alcohols, primary amines are oxidized to the corresponding imines, which can hydrolyze and react with unreacted amines (Fig. 16.7-1 B). In contrast to ethers, internal C-N bonds are readily oxidized, yielding substituted imines. This can be exploited for the production of substituted pyridines (Fig. 16.7-1 C). Furthermore, pyridines can be oxidized not only to N-oxides but also to a-hydroxylated products (Fig. 16.7-1 D). 

Use of a chiral proton source, a chiral base or base/chiral ligand complex circumvents the problem of incorporation and removal of a chiral auxiliary. Simpkins and coworkers opened the possibility of enantioselective protonation as a method for the asymmetric syntheses of 1-substituted tetrahydroisoquinolines [77]. Using the chiral amine 98 as a proton source, deracemization of 97 proceeded in up to 93 7 er, alleviating the requirement for a chiral auxiliary (Scheme 28). 

The synthesis of optically pure L-phenylglycine via the deracemization of mandelic acid was reported via three steps (racemization, enantioselective oxidation and stereoselective reductive amination). Racemization by mandelate racemase combined with simultaneous oxidation and reduction reactions with cofactor recycling gave the amino acid in 97% ee and 94% yield (Scheme 4.43) [96]. 

A common method for the deracemization of a-amino acids has been to employ amino acid oxidases along with a non-selective reduction of the intermediate imine by hydride-reducing agents (e.g., sodium boro hydride or sodium cyanoborohydride) or amine boranes [99, 100]. 

Scheme 4.46 One-pot, two-step deracemization of a-chiral primary amines.

---