Hostname: page-component-78c5997874-j824f Total loading time: 0 Render date: 2024-11-05T08:25:36.785Z Has data issue: false hasContentIssue false
  • English
  • Français

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]
Get access Rights & Permissions [Opens in a new window]

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

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Casal, E, Montilla, A, Moreno, FJ, Olano, A & Corzo, N 2006 Use of chitosan for selective removal of beta lactoglobulin from whey. Journal of Dairy Science 89 1384–1389CrossRefGoogle ScholarPubMed
Chiu, CK & Etzel, MR 1997 Fractionation of lactoperoxidase and lactoferrin from bovine whey using a cation exchange membrane. Journal of Food Science 62 996–1000CrossRefGoogle Scholar
De Wit, JN & Van Hooydonk, ACM 1996 Structure, functions and applications of lactoperoxidase in natural antimicrobial systems. Netherlands Milk and Dairy Journal 50 227–244Google Scholar
Foley, AA & Bates, GW 1987 The Purification of lactoferrin from human whey by batch extraction. Analytical Biochemistry 162 296–300CrossRefGoogle ScholarPubMed
Fonteh, FA 2001 Role of the lactoperoxidase system in raw milk preservation, PhD Thesis, University of ReadingGoogle Scholar
Fonteh, FA, Grandison, AS & Lewis, MJ 2002 Variations of lactoperoxidase activity and thiocyanate content in cows' and goats' milk throughout lactation. Journal of Dairy Research 69 401–409CrossRefGoogle ScholarPubMed
Goodall, S, Grandison, AS, Jauregi, P & Price, J 2008 Selective separation of the major whey proteins using ion exchange membranes. Journal of Dairy Science 91 1–10CrossRefGoogle ScholarPubMed
Hernández, MCM, Markwijik, V & Vreeman, HJ 1990 Isolation and properties of lactoperoxidase from bovine milk. Netherlands Milk and Dairy Journal 44 213–231Google Scholar
Morrison, M, Hamilton, HB & Stotz, E 1957 The isolation and purification of lactoperoxidase by ion exchange chromatography. Journal of Biological Chemistry 228 767–776CrossRefGoogle ScholarPubMed
Paul, KGH, Ohlsson, PI & Henriksson, A 1980 The Isolation and some ligandic properties of lactoperoxidase. FEBS Letters 110 200–204CrossRefGoogle ScholarPubMed
Polis, BD & Shmukler, HN 1953 Crystalline lactoperoxidase. Isolation by displacement chromatography. Journal of Biological Chemistry 201 475–500CrossRefGoogle ScholarPubMed
Rombatus, WA, Schroeder, WA & Morrison, M 1967 Bovine lactoperoxidase. Partial characterisation of the further purified protein. Biochemistry 6 2965–2977CrossRefGoogle Scholar
Uchida, T, Dosako, S, Sato, K & Kawakami, H 2003 Sequential separation of lactoferrin, lactoperoxidase and secretory components by sulfate-linked ion exchange chromatography. Milchwissenschaft 58 482–486Google Scholar
Yoshida, S 1988 Isolation of some minor proteins distributed in acid whey from approximately 100,000 to 250,000 Daltons of particle size. Journal of Dairy Science 71 1–9CrossRefGoogle Scholar
Yoshida, S & Ye-Xiuyun, 1991a Isolation of lactoperoxidase and lactoferrin from bovine milk rennet whey and acid whey by sulphopropyl cation-exchange chromatography. Netherlands Milk and Dairy Journal 45 273–280Google Scholar
Yoshida, S & Ye-Xiuyun, 1991b Isolation of lactoperoxidase and lactoferrin from bovine milk acid whey by carboxymethyl cation exchange chromatography. Journal of Dairy Science 74 1439–1444CrossRefGoogle Scholar