Unspecific peroxygenase immobilization in 3D-printed microfluidics: towards tailor-made screening platforms†

In the context of empowering biocatalysis, an easy-to-develop, reproducible, and easy-to-scale biocatalytic system under continuous flow is demonstrated. 3D printing technology is used as the reactor manufacturing method, yielding identical and low-cost microfluidic chips that can be further modified to serve as a biocatalytic platform for simultaneous parameter screening. The model enzyme studied here is unspecific peroxygenase (UPO). UPOs are currently under intensive study, due to their distinct promiscuity in oxyfunctionalization chemistry. This is the first study demonstrating UPO's immobilization in a microfluidic concept. The developed method for surface functionalization of microfluidic reactors is based on polydopamine modification and was proven highly reproducible. UPO showed a TTN of 19 249 and a STY of 2.1 g L⁻¹ h⁻¹, under the specified conditions. The kinetic behavior of the system under flow conditions is reported. The system was also regenerated with a 51.4% recovered activity. Further utilization of microfluidic concepts is expected to unravel the full potential of UPOs for oxyfunctionalization reactions of particular interest. The proposed system is foreseen as a screening platform for different reaction conditions, reaction substrates, or enzyme mutants.