Applied Biochemistry and Biotechnology 2010-02-01

Extraction of ascorbate oxidase from Cucurbita maxima by continuous process in perforated rotating disc contactor using aqueous two-phase systems.

T S Porto, P P Marques, C S Porto, K A Moreira, J L Lima-Filho, A Converti, A Pessoa, A L F Porto

Index: Appl. Biochem. Biotechnol. 160(4) , 1057-64, (2010)

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Abstract

The ascorbate oxidase is the enzyme used to determine the content of ascorbic acid in the pharmaceutical and food industries and clinics analyses. The techniques currently used for the purification of this enzyme raise its production cost. Thus, the development of alternative processes and with the potential to reduce costs is interesting. The application of aqueous two-phase system is proposed as an alternative to purification because it enables good separation of biomolecules. The objective of this study was to determine the conditions to continuously pre-purify the enzyme ascorbate oxidase by an aqueous two-phase system (PEG/citrate) using rotating column provided with perforated discs. Under the best conditions (20,000 g/mol PEG molar mass, 10% PEG concentration, and 25% citrate concentration), the system showed satisfactory results (partition coefficient, 3.35; separation efficiency, 54.98%; and purification factor, 1.46) and proved suitable for the pre-purification of ascorbate oxidase in continuous process.