Partition of horseradish peroxidase with maintained activity in aqueous biphasic system based on ionic liquid

Enzyme activity and partition behavior in aqueous biphasic systems (ABSs) consisting of ionic liquid (IL) and salt (IL-ABSs) were investigated to increase our understanding of IL-ABSs and shed light on their application potential as enzyme extraction system. With horseradish peroxidase (HRP) as the model enzyme, several effects of alkylimidazolium chloride–K2HPO4 ABSs on activity and partition behavior of enzyme were studied including alkyl chain length of ILs and concentrations of each component. High lyotropic ILs (1-butyl-3-methylimidazolium and 1-ethyl-3-methylimidazolium chloride) and adequate water content (>40%) were both essential for the activity maintenance of HRP in IL-ABS. 1-Butyl-3-methylimidazolium chloride ([C4mim]Cl) was found to be an appropriate IL for phase forming and HRP activity retaining. After optimization of phase condition, about 80% HRP amount was distributed in the IL-rich upper phase, and greater than 90% enzyme activity was obtained. Moreover, compared with the commonly used polymer-based ABSs, this [C4mim]Cl-ABS has a much lower viscosity, which is very beneficial to the experimental operation. Therefore, the tested IL-ABS could be considered as a potential enzyme extraction system.