Implications from protein adsorption onto anion- and cation-exchangers derivatized by modification of poly(ethylenimine)-Sepharose FF with succinic anhydride

Poly(ethylenimine) (PEI)-grafted Sepharose FF with an ionic capacity (IC) of 740 mmol/L (FF-PEI-L740) was modified with succinic anhydride, and six mix-charged resins of different ionic capacities (ICs) were synthesized [denoted as “Am-Cn” according to their anion exchange groups (amino groups) and cation exchange groups (carboxyl groups)]. Protein adsorption behaviors were investigated using bovine serum albumin (BSA) and lysozyme (LZM) as model proteins. The modification led to the net positive charge decrease to approximately zero (A320-C270), and then reversed the net charge to negative till the maximum negatively charge resin A20-C970. Adsorption capacity for BSA decreased from 205 ± 5 mg/mL (FF-PEI-L740) to 65 ± 3 mg/mL (A320-C270), but increased, for LZM, from 31 ± 1 mg/mL (A320-C270) to 281 ± 13 mg/mL (A20-C970) with increasing net negative charge. The uptake rate of BSA or LZM decreased with decreasing net positive charge or increasing net negative charge due to the decreased flexibility of the PEI chains brought from the increased electrostatic attraction or repulsion and/or tight binding due to the high net charge density. Effect of ionic strength (IS) showed that the resins with high net charge densities exhibited high adsorption capacities at high IS. Besides, dynamic binding capacity values achieved as high as 55 mg/mL for BSA and 143 mg/mL for LZM. Chromatographic experiments demonstrated favorable elution of bound proteins at NaCl lower than 370 mmol/L.