Abstract: The membrane of the activated glycidyl methacrylate (GMA)-modified cellulose was prepared and packed into a column piece by piece to make a microreactor by immobilization of trypsin. The microreactor based on the membrane medium showed the advantages of being cheaper, mechanically strong and chemically stable. The activity of immobilized trypsin towards N-benzoyl-L-arginine ethyl ester(BAEE) was 17800 U/g dry membrane, and was 52 % of that for free enzyme. Besides, the effects of pHvalue of buffer, temperature, ionic strength, organic modifier, and protein denaturants on the activity of immobilized trypsin were investigated in comparison to the free trypsin, and thermal stability also was found especially to be improved after immobilization. The activity of the immobilized trypsin showed no decay after continuously pumping BAEEthrough immobilized trypsin microreactor for 24 h at 40 ℃ and 0.5 mL/min. Finally, the cytochrome Cwas digested by microreactor and the products were analysed by MALDI-TOF-MS. The peptide mapping for a protein by combination of the immobilized enzyme membrane microreactor and MALDI-TOF-MShas been developed.

Key words: Glycidyl methacrylate (GMA)-modified cellulose membrane, Immobilized trypsin enzyme, Biological reactor, Degradation of protein, MALDI-TOF-MS, Cytochrome C