Rapid, Heterogeneous Biocatalytic Hydrogenation and Deuteration in a Continuous Flow Reactor

Abstract

The high selectivity of biocatalysis offers a valuable method for greener, more efficient production of enantiopure molecules. Operating immobilised enzymes in flow reactors can improve the productivity and handling of biocatalysts, and using H2 gas to drive redox enzymes bridges the gap to more traditional metal‐catalysed hydrogenation chemistry. Herein, we describe examples of H2‐driven heterogeneous biocatalysis in flow employing enzymes immobilised on a carbon nanotube column, achieving near‐quantitative conversion in <5 min residence time. Cofactor recycling is carried out in‐situ using H2 gas as a clean reductant, in a completely atom‐efficient process. The flow system is demonstrated for cofactor conversion, reductive amination and ketone reduction, and then extended to biocatalytic deuteration for the selective production of isotopically labelled chemicals.

Synthesis of [4S‐2H]NADH, [4R‐2H]NADH, [4‐2H2]NADH and [4‐2H]NAD+ cofactors through heterogeneous biocatalysis in heavy water

“…The main advantages over previous synthetic methods arise from the atom economy of using H 2 gas as a reductant and the simplicity of using 2 H 2 O as an isotope source, avoiding the requirement for carbon‐based co‐reagents. Furthermore, the inherent heterogeneous nature of the H 2 ‐driven biocatalyst makes reaction work‐up straightforward, and similar strategies have been demonstrated for translation into continuous flow 16 . Whilst the work presented here concerns cofactor deuteration, preliminary experiments in 3 H 2 O (37 MBq g −1 ) indicate that the same procedures should be translatable for similar tritiation.…”

Hybrid Chemo-, Bio-, and Electrocatalysis for Atom-Efficient Deuteration of Cofactors in Heavy Water

“…Given the lack of covalent attachment, the adsorption leads to surprisingly robust catalysts, and the systems show minimal enzyme leaching, even when they are operated under flow conditions. 32 …”

“…The hydrogenase and NAD + reductase can be electronically linked as part of a single enzyme or as distinct moieties immobilized on a conductive carbon support. 4 , 32 The separation of the two catalytic sites enables a 2 H-labeled product ([4- 2 H]-NADH) to be formed in the presence of an unlabeled reductant (H 2 ). In the work presented here, we seek to further establish this site-separated approach by comparing electrocatalytic, chemocatalytic, and biohybrid systems.…”

Accelerating Biocatalytic Hydrogenations using the H-Cube Flow Reactor

“…Such H2powered biotransformations have been demonstrated in batch reactions, and more recently have been extended to re-circulating flow reactors, 17 and into continuous flow. 3,16 In our previous work, biocatalytic hydrogenations were achieved using in house flow columns, but we were not able to achieve the control of H2 introduction that commercial reactors can provide. Here, we sought to demonstrate biocatalytic hydrogenation in an industry standard flow reactor, the H-Cube, Figure 1(d), for generation of reduced cofactor, NADH, and for NADH supply to NADH-dependent enzymes for C=O reductions.…”