The origin and consequences of concentration dependence in gel chromatography

Concentration dependence of elution volume was determined for Blue Dextran 2000, Dextran 500, Dextran sulphate 500 and bovine serum albumin on columns of Sephadex G-100 equilibrated with sodium phosphate buffer, I 0.1, pH6.8. From the results for Dextran 500, it was shown that a linear relation exists between elution volume and the corresponding osmotic pressure calculated for the same concentration and incorporating the term containing the second virial coefficient. This relationship was used to predict the concentration dependence of elution volume for bovine serum albumin and myoglobin, proteins that partially penetrate Sephadex G-100. Possible consequences of osmotic effects are considered in relation to various types of column experiments, including differential chromatography.