Immobilization of carbonic anhydrase on polyethylenimine/dopamine co-deposited SiO₂

Silica was modified by a polyethyleneimine (PEI)/dopamine (DA) co-deposition method, and then carbonic anhydrase (CA) was covalently immobilized via glutaraldehyde (GA). First, the effects of the mass ratio of PEI and DA, deposition time on the deposition rate were investigated, and silica microspheres before and after modification were characterized by Fourier transform infrared spectroscopy (FTIR) and scanning electron microscope (SEM). Then, the influences of the immobilization conditions including the deposition rate, the addition amount of PEI/PDA-SiO₂, CA and GA concentration on the activity recovery of immobilized enzyme were studied. At last, the storage stability and reusability of immobilized enzyme were also examined. The results show that the deposition rate increased firstly and then decreased with the mass ratio of PEI and DA increased, and reached the maximum at a mass ratio of 1:1. The deposition rate increased linearly with the time prolonged, which respectively reached 1.33% and 0.5% after 10 h for PEI/DA co-deposited system and pure DA-deposited that. The deposition rate had no obvious effect on the content of N element and activity recovery of immobilized CA. For enzyme immobilization, there was a saturated addition amount of the carrier which was 0.25 g PEI/PDA-SiO₂/mg CA, and the optimal concentration of CA and GA was 0.8 mg/ml and 0.1%(mass), respectively. Under these conditions, the activity recovery of CA-PEI/PDA-SiO₂ was about 78.8%. After 30 days of storage at 25℃, the remaining activity of immobilized CA was 77.2% while that of free CA was only 12%. The relative activity of immobilized CA was 88.3% after undergoing 10 cycles.