Improved purification of β-lactoglobulin from acid whey by means of ceramic hydroxyapatite chromatography with sodium fluoride as a displacer

The successful separation of β-lactoglobulin from other bovine whey proteins was performed by ceramic hydroxyapatite chromatography with a fluoride ion gradient in phosphate buffer as displacement agent. The method was applied to acid whey originating from milk of healthy as well as of mastitic cows. β-Lactoglobulin was completely eluted in one peak at a fluoride concentration of about 0.6 mol/l. The purity of β-lactoglobulin in this fraction was at least 96% if whey from healthy milk was processed. Co-eluted contaminants are traces of immunoglobulin G, serum albumin and lactoferrin. In case of mastitic whey the proportion of β-lactoglobulin is diminished as the amounts of immunglobulin G, serum albumin and lactoferrin are increased within this fraction. Size exclusion chromatography on Superdex 75 pg effectively removed contaminants resulting in a purity for β-lactoglobulin from normal whey of approximately 99%. The yield of β-lactoglobulin from physiological whey was 50–55% referring to the fraction highly enriched with β-lactoglobulin by hydroxyapatite chromatography. In case of mastitic milk the higher amounts of contaminants were also removed successfully by size exclusion chromatography.