Fractionation of whey proteins with high-capacity superparamagnetic ion-exchangers.
In this study we describe the design, preparation and testing of superparamagnetic anion-exchangers, and their use together with cation-exchangers in the fractionation of bovine whey proteins as a model study for high-gradient magnetic fishing. Adsorbents prepared by attachment of trimethyl amine to particles activated in sequential reactions with allyl bromide and N-bromosuccinimide yielded a maximum bovine serum albumin binding capacity of 156 mg g(-1) combined with a dissociation constant of 0.60 microM, whereas ion-exchangers created by linking polyethylene imine through superficial aldehydes bound up to 337 mg g(-1) with a dissociation constant of 0.042 microM. The latter anion-exchanger was selected for studies of whey protein fractionation. In these, crude bovine whey was treated with a superparamagnetic cation-exchanger to adsorb basic protein species, and the supernatant arising from this treatment was then contacted with the anion-exchanger. For both adsorbent classes of ion-exchanger, desorption selectivity was subsequently studied by sequentially increasing the concentration of NaCl in the elution buffer. In the initial cation-exchange step quantitative removal of lactoferrin (LF) and lactoperoxidase (LPO) was achieved with some simultaneous binding of immunoglobulins (Ig). The immunoglobulins were separated from the other two proteins by desorbing with a low concentration of NaCl (< or = 0.4 M), whereas lactoferrin and lactoperoxidase were co-eluted in significantly purer form, e.g. lactoperoxidase was purified 28-fold over the starting material, when the NaCl concentration was increased to 0.4-1 M. The anion-exchanger adsorbed beta-lactoglobulin (beta-LG) selectively allowing separation from the remaining protein.
Heebøll-Nielsen
Anders
A
Center for Process Biotechnology, BioCentrum-DTU, Technical University of Denmark, DK-2800, Kgs. Lyngby.
Justesen
Sune F L
SF
Thomas
Owen R T
OR
D000327:Adsorption; D000818:Animals; D002417:Cattle; D002611:Cheese; D005591:Chemical Fractionation / Q000295:instrumentation / Q000379:methods*; D002852:Chromatography, Ion Exchange; D004983:Ethanolamines; D019649:Food Industry / Q000295:instrumentation / Q000379:methods*; D007700:Kinetics; D008280:Magnetics; D008744:Methylamines; D008894:Milk Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000493:pharmacokinetics; D011094:Polyethyleneimine; D011485:Protein Binding; D000067816:Whey Proteins
D004983:Ethanolamines; D008744:Methylamines; D008894:Milk Proteins; D000067816:Whey Proteins; D011094:Polyethyleneimine; C023336:trimethylamine; C007369:2-diethylaminoethanol