Key role of dialysis membranes for the enzyme utilization and reactor productivity in sustainable aldol synthesis of l-phenylserine

The rapid deactivation of free enzymes, often occurring at liquid-liquid interfaces, represents a significant challenge limiting the commercialization of many enzymatic reactions. Potential solutions, such as enzyme or reaction mixture modifications, typically need to be tailored to each specific reaction system. Here, we propose a universal approach to protect the enzyme from direct contact with liquid-liquid interfaces. This approach employs dialysis membranes, which create a solid but permeable interface between aqueous and organic phases. We tested this concept, implemented into both batch and semi-continuous milli-reactors, on aldol synthesis of l-phenylserine (LPS) diastereoisomers using an enzyme l-threonine aldolase in an aqueous solution, with glycine and benzaldehyde as substrates. Integrating a dialysis membrane into a semi-continuous milli-reactor preserved enzyme activity for over three days, in contrast to rapid deactivation observed when the aqueous reaction mixture was in direct contact with the benzaldehyde phase. Furthermore, employing a dialysis membrane in the batch reactor led to a substantial increase in LPS concentration (101 mM vs. 64 mM) and glycine utilization (14.5% vs. 8.5%) compared to systems with direct phase contact. The choice between a batch or semi-continuous membrane milli-reactor depends on specific requirements for enzyme stability and product concentration. Notably, the scalable semi-continuous membrane process provides industrially relevant productivity. These findings underscore the potential of membrane-assisted processes to facilitate scalable and sustainable enzymatic synthesis of chiral compounds.