Fractionation of Caseins Directly from Skimmilk by Gel Chromatography. 2. Elution with Phosphate Buffers

Abstract

n-Casein was separated on Sephadex G-100 by eluting with 0.01 ~ borate, pH 10 at 25 C or 0.01 M phosphate, pit 11 at 4 C. Phosphate, 5 m~ with 2 mM Na ethylenediaminetetraacetate, pH 10.8, was used for continuous fractionation at 4 C to retard bacterial growth in the gel. ~-Casein was collected from the first half of the first peak. The stabilizing ability o~ the K-casein against Ca-asl-caseinate precipitation was slightly low possibly from contamination with minor members of the a~-casein group. Electrophoretically pure t-casein was obtained from the first half of the second peak after elimination of contaminating a~-easein by precipitation at pH 4.4 and 4 C. a~-Cascin was purified from the last half of the second peak by precipitating with 0.1 ~ CaClo. The phosphate method was superior to the sodium dodecylsulfate method because of faster elution, being more reproducible, and without extraction to remove bound dodecylsulfate, a~-, fl-, n-Caseins were fractionated directly from skimnfilk, however, the yield of n-casein was approximately half of that by the sodium dodecylsulfate method.