Optimised Protocol for Drying Aqueous Enzyme Solutions in Organic Solvents - Comparison of Free and Immobilised Candida antarctica Lipase B

Here, we propose an optimised protocol for controlling the initial water activity (a(w)) in organic reaction mixtures with soluble lipase to ensure reproducible and consistent enzyme activity measurements. Pre-equilibration above saturated salt solutions was used to set a(w) of the reaction media, where a(w) was tracked with a hygrometer. Consistent stirring of the suspension and volume correction to compensate for solvent evaporation were found to be critical. The protocol was tested on Candida antarctica lipase B (CALB) in soluble and immobilised forms in three organic solvents (acetonitrile, methyl tert-butyl ether, and hexane) at four different water activities ranging from 0.12 to 0.97. Soluble and immobilised CALB had similar a(w)-profiles and showed the highest enzyme activity in hexane. The optimised protocol expands the possibility to study enzyme kinetics from immobilised enzymes to soluble enzymes.