Lipase from Pseudomonas fluorescens (Amano AK) has been immobilized by the sol–gel method using tetramethoxysilane and trimethoxysilanes with alkyl or aryl groups as precursors and ionic liquids as immobilization additives. Room temperature ionic liquids (ILs) based on imidazolium cations with different hydrophobicities and inorganic or organic anions were investigated. The biocatalytic activity in the acylation reaction of secondary aliphatic alcohols by vinyl acetate increased at lower polarity of the IL, being 58% higher in case of octyl than for ethyl substituent of the alkylimidazolium cation. The optimum IL/silane molar ratio in the immobilization mixture was around 0.2. Hydrophobic groups from both the silane precursor and IL converge to the optimal microenvironment for the enzyme action. ILs as reaction media led to higher activities as common organic solvents in the kinetic resolution of secondary alcohols, but the highest enantioselectivities (enantiomeric ratio E values > 50) have been observed in acetone and tetrahydrofuran. Structural investigation of the preparates by SEM–EDX and FT-IR has confirmed the partial confinement of the IL in the xerogel structure.