Effects of bovine serum albumin heterogeneity on frontal analysis with anion-exchange media

The presence of dimers in commercial bovine serum albumin (BSA) samples of nominal high purity is investigated along with their effects on the frontal analysis behavior of preparative anion-exchange media. Size-exclusion and analytical anion-exchange chromatography are used to determine the relative amounts of monomer and dimer for two samples of BSA. While the amount of dimer was generally low, its impact on adsorptive behavior is significant. Equilibrium experiments demonstrate that the anion-exchange media binds the dimer more strongly, leading to an unequal distribution of the monomer and dimer between the two phases. Analysis of the breakthrough behavior of BSA reveals that the monomer breaks through prior to the dimer. This leads to the characteristic tailing breakthrough curve often seen with BSA. Breakthrough experiments carried out using dimer-free BSA confirm that the extreme tailing observed with the commercial samples is curtailed by removing the dimer.