Enhancement of the excluded-volume effect in protein extraction using triblock copolymer-based aqueous micellar two-phase systems

Triblock copolymer surfactants consisting of poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO), Pluronic L61 (PEO-PPO-PEO, L61) and Pluronic 25R2 (PPO-PEO-PPO, 25R2) were exploited in aqueous micellar two-phase syste ms for the protein extraction. The extraction was based on the phase separa tion into surfactant-depleted and -condensed phases (an aqueous and a surfa ctant-rich phases, respectively) upon warming aqueous micellar solutions of triblock copolymer. In both systems, hydrophilic proteins such as albumin were not extracted into the surfactant-rich phase. On the other hand, hydro phobic cytochrome b(5) was well extracted in the L61 system due to hydropho bic interaction. However, the extraction of cytochrome b(5) was not observe d in the 25R2 system. This abnormal extractability of cytochrome b5 in the 25R2 system was explained by the enhanced excluded-volume interaction betwe en cytochrome b(5) and 25R2 micellar network in the surfactant-rich phase, which overcomes the hydrophobic interaction. Additionally, ionic surfactant s were added into the systems for controlling extractability of proteins. I n the 25R2 system, cationic tetradecyltrimethylammonium was effective for e xtracting anionic cytochrome bs against the excluded-volume effect, while n ot for anionic albumin because of its large molecular weight. In 25R2 syste m containing ionic surfactant, the partitioning of proteins were found to b e governed by the hydrophobic, excluded-volume, and electrostatic interacti ons. Micellar network formed by 25R2 type of surfactant with a strong exclu ded-volume interaction could provide new selective extraction systems for t he separation of proteins. (C) 2001 Elsevier Science B.V. All rights reserv ed.