Recovery of Peniophora cinerea laccase using aqueous two-phase systems composed by ethylene oxide/propylene oxide copolymer and potassium phosphate salts.
Aqueous two-phase systems (ATPSs) composed by UCON (ethylene oxide/propylene oxide copolymer) and potassium phosphate salts were for the first time evaluated in the recovery of Peniophora cinerea laccase from complex fermented medium. The ATPSs were obtained by combining the random copolymer UCON with KH2PO4, potassium phosphate buffer pH 7 or K2HPO4. According to the results, protein partition occurred predominantly toward the saline phase (bottom phase) of the ATPSs, while some contaminants such as pigments partitioned mainly to the top phase. In preliminary tests, it was found that the salt with the lowest pH value (KH2PO4, pH 4.6) stimulated the enzyme activity, while the other salts (pH between 7.0 and 9.5) caused a strong inhibition. However, the salt inhibition was not observed in the equilibrium phases of the UCON-Potassium phosphate ATPSs. The laccase recovery was high for all the biphasic systems, but the highest value (134%) was obtained when using UCON combined with KH2PO4. When compared to conventional concentration and purification methods (lyophilization, ammonium sulfate precipitation, ultrafiltration, and ion exchange chromatography), ATPS was demonstrated to be an efficient alternative for P. cinerea laccase recovery from fermented medium.
Department of Biotechnology, Engineering College of Lorena, University of São Paulo, Estrada Municipal do Campinho s/n, CEP, 12602-810 Lorena/SP, Brazil.
Sérgio
S
Moreira
Sara C
SC
Silvério
Eugénia A
EA
Macedo
Adriane M F
AM
Milagres
José A
JA
Teixeira
Solange I
SI
Mussatto
D001487:Basidiomycota / Q000201:enzymology*; D004852:Epoxy Compounds / Q000737:chemistry*; D005027:Ethylene Oxide / Q000737:chemistry*; D042845:Laccase / Q000302:isolation & purification*; D010710:Phosphates / Q000737:chemistry*; D017680:Potassium Compounds / Q000737:chemistry*; D012492:Salts
D004852:Epoxy Compounds; D010710:Phosphates; D017680:Potassium Compounds; D012492:Salts; C013216:potassium phosphate; D042845:Laccase; D005027:Ethylene Oxide; C009068:propylene oxide
Aqueous two-phase systems; Laccase; Phosphate salts; Purification; UCON