Separation of lactoferrin from whey by dye affinity chromatography

M. GRASSELLI; O. CASCONE

Netherlands Milk Dairy Journal

Association for the avancement of dairy science

Año: 1996 vol. 50 p. 551 - 561

0028-209X

Eight triazinic dyes were assayed as ligands for chromatographic affinity purification of lactoferrin from rennet whey, at pH 5.0, 7.0 and 9.0. Blue R-HE, Orange R-HE and Scarlet G-A did not adsorb lactoferrin at any pH, whereas Ye1low FR adsorbed a small amount of lactoferrin and showed poor selectivity. Red HE-3B was most selective and lactoferrin was most adsorbed at pH 5.0 and 7.0. Blue F3-GA, Red HE-7B and Red F-5B adsorbed intermediate amounts of lactoferrin. At pH 9.0, Blue F3-GA, Red HE-3B and Red F-5B adsorbed intermediate amounts of lactoferrin. Red HE-7B maintained its low lactoferrin adsorption and good selectivity. At pH 5.0-7.0, lactoferrin shows high adsorption on Red HE-3B and at pH 7.0, selectivity was better than at pH 5.0, because lactoferrin was accompanied by only a small amount of B-lactoglobulin. Red HE-3B was selected for tests on ionic-strength gradient elution. At pH 7.0, lactoferrin eluted with NaCl at about 1 mol/L, and the contaminating B-lactoglobulin did so with NaCl at 0.3 mol/L. Treatment of immobilized Red HE-3B with sodium dithionite and sodium nitrite did not improve separation and considerably decreased recovery. A technique was developed for batch purification based on data from gradient elution. The procedure allowed extraction of 82% of the lactoferrin whey content with a purity of 98%.