ETHANOL SODIUM-CHLORIDE PHOSPHATE MOBILE-PHASE FOR SIZE-EXCLUSION CHROMATOGRAPHY OF POLY(ETHYLENE GLYCOL) MODIFIED PROTEINS

The use of an aqueous sodium chloride-phosphate mobile phase for size- exclusion chromatography of poly(ethylene glycol) modified (PEGylated) proteins resulted in premature degradation of silica-based gel filtra tion columns. This column degradation was manifested as peak tailing a nd loss of resolution. An aqueous ethanol-sodium chloride-phosphate mo bile phase was identified which extended column lifetimes without loss of resolution or change in peak shape. We describe the effects of sod ium chloride, phosphate, and ethanol concentrations on column performa nce. Peak shape and column lifetimes were better when an aqueous ethan ol-sodium chloride-phosphate mobile phase was used than with an aqueou s salt mobile phase. Also, circular dichroism was utilized to investig ate possible solvent effects on the conformation of each protein.