Effective digestion of casein with agar gel-entrapped cell wall fraction-bound enzymes of Aspergillus oryzae as immobilized proteases

Abstract Multienzyme hydrolysis of casein by an immobilized biocatalyst was investigated using cell wall fraction-bound enzymes (CWEs) in Aspergillus oryzae . The bioreactor was made up of a CWE system entrapped into agar gel. Hydrolyzates of casein were analyzed by a modified Lowry, Ninhydrin, and o -phthaldialdehyde (OPA) methods, SDS-PAGE and HPLC. The degree of hydrolysis became maximum after 48 h-reaction. From the molecular weight of hydrolyzates, 59%, 95% and 100% of the original casein was digested after 12 h-, 24 h- and 48 h-reaction, respectively. The increase of peptides (200–2000 in molecular weight) was especially remarkable. Yet the activity of the immobilized CWEs remained at about 50% of the initial activity after 9 times repeated use. Our results suggest that the immobilized CWE system is a new type of immobilized biocatalyst possessing the characteristics of both immobilized enzymes and immobilized viable cells. This suggests the possibility that the drawbacks of immobilized enzymes against high molecular weight substrates can be improved.