During the downstream processes of protein biologics, the protein monomer must be separated from the aggregates. This is usually carried out by using electrostatic interaction chromatography. However, since it is not an efficient process, the process optimization is needed, which is not an easy task.<br/>One of the attractive methods for improving the separation performance of a monomer from the aggregates is to add PEG into the mobile phase (Method A). The peak retention volume shifts to larger volumes in the presence of PEG. The shift increases with the protein molecular weight.<br/>Another method is to use a different type of ion-exchange ligand. We have shown that the poly-amine based anion-exchange ligand is effective for monomer-dimer separations (Method B).<br/>In this study we have compared the performance of monomer-dimer separations by the above-mentioned two methods. One disadvantage of Method A is an increased viscosity due to PEG, which results in higher back pressures and peak broadening. Another problem of this method is a low solubility of proteins near its pI. We have analyzed the peak broadening in terms of the pore diffusion coefficient. Method B is also known as a salt-tolerant chromatography, in which much higher salt concentrations are needed for elution. As proteins are retained even slightly below the pI with this column, the separation can be carried out at a relatively wide range of mobile phase pH.<br/>

Optimization of monomer separation processes by electrostatic-interaction chromatography

249th National Meeting (2015)

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