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Extraction of Ascorbate Oxidase from Cucurbita maxima by Continuous Process in Perforated Rotating Disc Contactor Using Aqueous Two-Phase Systems

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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.

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Acknowledgements

The authors wish to acknowledge the financial support of FAPESP (Fundação de Amparo à Pesquisa do Estado de São Paulo, São Paulo, Brazil), CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico, Brasilia, Brazil), and CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, Brasilia, Brazil).

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Authors and Affiliations

  1. Departament of Biochemical and Pharmaceutical Technology, University of Sao Paulo, Sao Paulo, Brazil

    T. S. Porto & A. Pessoa

  2. Laboratory of Immunopathology Keizo Asami (LIKA), University Federal of Pernambuco, Cidade Universitária, 50670-901, Recife, Pernambuco, Brazil

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

  3. Academic Unit of Garanhuns, University Federal Rural of Pernambuco, Recife, Brazil

    K. A. Moreira

  4. Department of Chemical and Process Engineering, University of Genoa, Genoa, Italy

    A. Converti

  5. Department of Animal Morphology and Physiology, University Federal Rural of Pernambuco, Recife, Brazil

    A. L. F. Porto

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  1. T. S. Porto
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  2. P. P. Marques
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  3. C. S. Porto
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  4. K. A. Moreira
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  5. J. L. Lima-Filho
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  6. A. Converti
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  7. A. Pessoa
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  8. A. L. F. Porto
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Correspondence to T. S. Porto.

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Porto, T.S., Marques, P.P., Porto, C.S. et al. Extraction of Ascorbate Oxidase from Cucurbita maxima by Continuous Process in Perforated Rotating Disc Contactor Using Aqueous Two-Phase Systems. Appl Biochem Biotechnol 160, 1057–1064 (2010). https://doi.org/10.1007/s12010-009-8631-y

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  • DOI: https://doi.org/10.1007/s12010-009-8631-y

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