A Millifluidic Device with Embedded Cross-Linked Enzyme Aggregates for Degradation of H2O2

Millifluidic devices decorated with enzymes have been used for enzymatic reactions in continuous processes, but low enzymatic activity and enzyme leaching remain as challenges. Herein, we develop a strategy to embed cross-linked enzyme aggregates (CLEAs) on the surfaces of millifluidic devices to achieve higher enzymatic activity and better stability. Catalase was chosen as a model enzyme to degrade H2O2 in wastewater samples. First, CLEA of catalase (153 +/- 10 nm) was formed by simultaneous precipitation and cross-linking with 25.0 wt % acetonitrile containing 0.025 wt % glutaraldehyde in a millifluidic device. To immobilize CLEA, we first swell a piece of plastic tubing by using 5.0 wt % acetonitrile and then immerse it in an aqueous solution with 5.0 wt % (3-aminopropyl)triethoxysilane (APTES) and 5.0 wt % dextran polyaldehyde (DPA) subsequently. After CLEA is absorbed inside the expanded polymer network of the tubing, the tubing is tightened by using a vacuum to secure the immobilized CLEA. The millifluidic device decorated with CLEA of catalase has total activity of 660 U for degradation of H2O2, and it shows good stability under a flow rate of 200 mu L/min. The tubing can be used to degrade 0.1 wt % H2O2 solution continuously for 3 h or remove 2 wt % residual H2O2 in wastewater for 2 h. The technique is general enough and can be applied to other types of enzymes for continuous enzymatic reactions.