title: Purification and in situ immobilization of papain with aqueous two-phase system.
abstract: Papain was purified from spray-dried Carica papaya latex using aqueous two-phase system (ATPS). Then it was recovered from PEG phase by in situ immobilization or preparing cross-linked enzyme aggregates (CLEAs). The Plackett-Burman design and the central composite design (CCD) together with the response surface methodology (RSM) were used to optimize the APTS processes. The highly purified papain (96-100%) was achieved under the optimized conditions: 40% (w/w) 15 mg/ml enzyme solution, 14.33-17.65% (w/w) PEG 6000, 14.27-14.42% (w/w) NaH2PO4/K2HPO4 and pH 5.77-6.30 at 20°C. An in situ enzyme immobilization approach, carried out by directly dispersing aminated supports and chitosan beads into the PEG phase, was investigated to recover papain, in which a high immobilization yield (>90%) and activity recovery (>40%) was obtained. Moreover, CLEAs were successfully used in recovering papain from PEG phase with a hydrolytic activity hundreds times higher than the carrier-bound immobilized papain.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, China.
authors.forename: Mingliang
authors.initials: M
authors.lastname: Li
authors.forename: Erzheng
authors.initials: E
authors.lastname: Su
authors.forename: Pengyong
authors.initials: P
authors.lastname: You
authors.forename: Xiangyu
authors.initials: X
authors.lastname: Gong
authors.forename: Ming
authors.initials: M
authors.lastname: Sun
authors.forename: Diansheng
authors.initials: D
authors.lastname: Xu
authors.forename: Dongzhi
authors.initials: D
authors.lastname: Wei
mesh_terms: D001671:Biochemistry / Q000379:methods; D029441:Carica / Q000378:metabolism*; D048271:Chitosan / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003432:Cross-Linking Reagents / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004798:Enzymes / Q000737:chemistry; D004800:Enzymes, Immobilized / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D007840:Latex / Q000737:chemistry; D010206:Papain / Q000737:chemistry* / Q000302:isolation & purification; D011092:Polyethylene Glycols / Q000737:chemistry; D013696:Temperature; D014867:Water / Q000737:chemistry
chemical_list: D003432:Cross-Linking Reagents; D004798:Enzymes; D004800:Enzymes, Immobilized; D007840:Latex; D014867:Water; D011092:Polyethylene Glycols; D048271:Chitosan; D010206:Papain