Immobilization of transaminases creates promising biocatalysts for production of chiral amines in batch or continuous-flow mode reactions. E. coli cells containing overexpressed transaminases of various selectivities and hollow silica microspheres as supporting agent were immobilized by an improved sol-gel process to produce immobilized transaminase biocatalysts with suitable stability and mechanical properties for continuous-flow applications. The immobilized cell-based transaminase biocatalyst proved to be durable and easy-to-use in kinetic resolution of four racemic amines 1a–d. The batch and continuous-flow mode kinetic resolutions with transaminase biocatalyst of opposite stereopreference provided access to both enantiomers of the corresponding amines. By using the most suitable immobilized transaminase biocatalysts, this study describes the first transaminase-based approach for the production of both pure enantiomers of 1-(3,4-dimethoxyphenyl)ethan-1-amine 1d.

This study implements a convenient microreactor for biocatalysis with enzymes immobilized on magnetic nanoparticles (MNPs). The enzyme immobilized onto MNPs by adsorption or by covalent bonds was lipase B from Candida antarctica (CaLB). The MNPs for adsorption were obtained by covering the magnetite core with a silica shell and later with hexadecyltrimethoxysilane, while for covalent immobilization, the silica-covered MNPs were functionalized by a layer forming from mixtures of hexadecyl- and 3-(2-aminoethylamino)propyldimethoxymethylsilanes in 16:1 molar ratio, which was further activated with neopentyl glycol diglycidyl ether (NGDE). The resulting CaLB-MNPs were tested in a convenient continuous flow system, created by 3D printing to hold six adjustable permanent magnets beneath a polytetrafluoroethylene tube (PTFE) to anchor the MNP biocatalyst inside the tube reactor. The anchored CaLB-MNPs formed reaction chambers in the tube for passing the fluid through and above the MNP biocatalysts, thus increasing the mixing during the fluid flow and resulting in enhanced activity of CaLB on MNPs. The enantiomer selective acylation of 4-(morpholin-4-yl)butan-2-ol (±)-1, being the chiral alcohol constituent of the mucolytic drug Fedrilate, was carried out by CaLB-MNPs in the U-shape reactor. The CaLB-MNPs in the U-shape reactor were compared in batch reactions to the lyophilized CaLB and to the CaLB-MNPs using the same reaction composition, and the same amounts of CaLB showed similar or higher activity in flow mode and superior activity as compared to the lyophilized powder form. The U-shape permanent magnet design represents a general and easy-to-access implementation of MNP-based flow microreactors, being useful for many biotransformations and reducing costly and time-consuming downstream processes.

As efforts in rational drug design are driving the pharmaceutical industry towards more complex molecules, the synthesis and production of these new drugs can benefit from new reaction routes. In addition to the introduction of new centers of asymmetry, complexity can be also increased by ring saturation, which also provides improved developability measures. Therefore, in this report, our aim was to develop transaminase (TA)-catalyzed asymmetric synthesis of a new group of potential chiral drug scaffolds comprising a saturated amine heterocycle backbone and an asymmetric primary amine sidechain (55a–g). We screened the Codex^® Amine Transaminase Kit of 24 transaminases with the morpholine containing ketone 57a, resulting in one (R)-selective TA and three (S)-selective TAs operating at 100 mM substrate concentration and 25 v/v% isopropylamine (IPA) content. The optimized reaction conditions were than applied for asymmetric transamination of further six ketones (57b–g) containing various amine heterocycles, in which a strong effect of the substitution pattern of the γ-position relative to the substituted N-atom could be observed. Mediated by the most enantiotope selective (S)-TAs in scaled-up process, the (S)-amines [(S)-55a–g] were isolated with moderate-to-excellent yields (47–94%) in enantiopure form (>99% ee).