Immobilization of cholesterol esterase in mesoporous silica materials and its hydrolytic activity toward diethyl phthalate

Cholesterol esterase (CE, cholesteryl ester hydrolase, EC 3.1.1.13) from porcine pancreas (molecular weight 400–500 kDa) exhibits hydrolytic activity toward various toxic organic phthalate esters. CE was confined in the nanospace (diameter 3–30 nm) of five types of mesoporous silica (MPS) that differ in structural properties such as pore diameter, pore volume, and particle morphology. These structural properties were characterized by transmission electron microscopy, small-angle X-ray diffraction, N2 adsorption–desorption experiments, solid-state 13C nuclear magnetic resonance (NMR), and solid-state 29Si NMR. Catalytic activities of immobilized and free CE were evaluated by the hydrolysis of diethyl phthalate in phosphate buffer solutions containing an organic cosolvent. Optimal activity recovery was achieved when CE was immobilized in n-decane-functionalized MPS, which had a large pore size (22.5 nm). The immobilization also protected against effects of temperature within the range 30 °C–60 °C; CE immobilized in n-decyl-functionalized MPS exhibited better thermal stability than in non-functionalized MPS or free CE. Moreover, it retained approximately 60% of its catalytic activity even after six catalytic cycles.

► The highest activity of immobilized CE was shown in MPS with a pore size of 22.5 nm. ► Catalytic efficiency improved when MPS was functionalized by n-decyl substitution. ► Immobilized CE exhibited good thermal stability and reusability. ► Organic co-solvent and the substrate structures affected enzyme activities.