High transesterification activities of immobilized proteases in new ether-functionalized ionic liquids

Three new ether-functionalized hydrophobic ionic liquids (ILs) were synthesized. Two proteases (subtilisin and α-chymotrypsin both covalently immobilized on chitosan) exhibited high synthetic activity (1–3 μmol/min·g) and selectivity (>97–99%, esterification over hydrolysis) in these ILs containing 10–15% (v/v) water during the transesterification of N-acetyl-l-phenylalanine ethyl ester with 1-propanol. However, the same reaction in t-butanol or [BMIM][Tf2N], with water contents higher than 2% (v/v) yielded low synthetic activities (0.2–1 μmol/min g in t-butanol) and/or poor selectivity (<40%). The high synthetic activities of proteases in ether-functionalized ILs at high water contents are explained by two reasons: (1) the hydrogen-bond donors in ILs (R1–O–R2) and chitosan (–OH) controlling the thermodynamic water activity of the reaction system, and (2) the protective role of the ether chain in reducing the cation–protein interaction.