Electrostatic effects in SEC of proteins

Electrostatic contributions are undoubtedly important in protein chromatography but these effects may be difficult to isolate. As we have seen, the ionic strength dependence of the elution volume is a key diagnostic of coulom-bic substrate-solute interactions. For proteins, however, hydrophobic interactions with the packing may comprise part of the effect of simple electrolyte. The interpretation of pH effects is also more complex for proteins than for strong polyelectrolytes, because in the former case the charge density on the 

Studying the behavior of bovine serum albumin (BSA) and acetycholinestrase (AChE) on untreated CPG in pH 8 buffer, Crone (29) observed separations only at ionic strengths near 0.05 M for I 0.01 M, only exclusion peaks were found, while chromatography at I 0.15 M produced partial or total adsorption. Pre-treatment of the column with PEO reduced adsorption and so more clearly revealed the electrostatic repulsion between AChE and the packing Ksec was found to increase from 0.13 to 0.52 as I increased from 0.0025 M to 0.15 M. 

Untreated CPG has also been used to fractionate casein micelles from milk (70). In order to completely avoid the adsorption of proteins, however, PEO pre-treatment has been employed (71). In this way, size separations are obtained in 0.05 ionic strength eluant, but with poor efficiency. 

---