Co-Confinement of Enzymes and Cofactors Within Pickering Droplet Derived Microreactors for Continuous Flow Catalysis

The co-immobilization of enzymes and cofactors represents a sustainable platform for continuous-flow synthesis of chiral pharmaceuticals, yet balancing effective retention with mutual accessibility of them remains challenging. Herein, we report a sol-gel strategy to construct Pickering droplet derived microreactors (PDMRs) for the co-confinement of enzymes and cofactors, which have been applied to continuous flow reactions without exogenous addition of cofactors. Within these self-sufficient PDMRs, the cofactors are reversibly immobilized via electrostatic interactions, enabling in situ regeneration and free access to the enzyme. The PDMRs efficiently encapsulate enzymes and cofactors with 85-100% immobilization efficiency and robust thermal stability. In PDMRs-catalyzed continuous flow reactions, excellent catalytic performance and high cofactor total turnover number (TTN) were obtained in nicotinamide adenine dinucleotide phosphate (NADP⁺)-dependent aldo-keto reductase (AKR) catalyzed enantioselective reductions (80-100% conversions, >99% ee, 500 h stability, up to 173 907 mol mol^-1 TTN), and pyridoxal 5-phosphate (PLP)-dependent transaminase (TA) catalyzed enantioselective transaminations (80-100% conversions, >99% ee, 2 000 h stability, up to 45 552 mol mol^-1 TTN). Furthermore, the PDMRs are extended to the co-confinement of a multi-enzyme system (AKR and glucose dehydrogenase, GDH) with NADP⁺ for chiral alcohols synthesis with sustained operational stability. This work establishes a potent and durable strategy for industrial-scale continuous flow manufacturing.