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Isolation of lactoperoxidase using different cation exchange resins by batch and column procedures

Published online by Cambridge University Press:  19 May 2010

Leonard WT Fweja ,
Michael J Lewis  and
Alistair S Grandison
Leonard WT Fweja
Affiliation:
Department of Food and Nutritional Sciences, University of Reading, P O Box 226, Whiteknights, Reading, RG6 6AP, UK
Michael J Lewis
Affiliation:
Department of Food and Nutritional Sciences, University of Reading, P O Box 226, Whiteknights, Reading, RG6 6AP, UK
Alistair S Grandison*
Affiliation:
Department of Food and Nutritional Sciences, University of Reading, P O Box 226, Whiteknights, Reading, RG6 6AP, UK
*
*For correspondence; e-mail: [email protected]
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Abstract

Lactoperoxidase (LP) was isolated from whey protein by cation-exchange using Carboxymethyl resin (CM-25C) and Sulphopropyl Toyopearl resin (SP-650C). Both batch and column procedures were employed and the adsorption capacities and extraction efficiencies were compared. The resin bed volume to whey volume ratios were 0·96:1·0 for CM-25C and ⩽0·64:1·0 for SP-650 indicating higher adsorption capacity of SP-650 compared with CM-25C. The effluent LP activity depended on both the enzyme activity in the whey and the amount of whey loaded on the column within the saturation limits of the resin. The percentage recovery was high below the saturation point and fell off rapidly with over-saturation. While effective recovery was achieved with column extraction procedures, the recovery was poor in batch procedures. The whey-resin contact time had little impact on the enzyme adsorption. SDS PAGE and HPLC analyses were also carried out, the purity was examined and the proteins characterised in terms of molecular weights. Reversed phase HPLC provided clear distinction of the LP and lactoferrin (LF) peaks. The enzyme purity was higher in column effluents compared with batch effluents, judged on the basis of the clarity of the gel bands and the resolved peaks in HPLC chromatograms.

Keywords

LactoperoxidaseCarboxymethyl resinSulphopropyl Toyopearl resincation-exchange
Type
Research Article
Information
Journal of Dairy Research , Volume 77 , Issue 3 , August 2010 , pp. 357 - 367
Copyright
Copyright © Proprietors of Journal of Dairy Research 2010

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