Extraction of Proteins with Aqueous Two-Phase Systems Formed by Protic Ionic Liquids and Inorganic Salts

Abstract

A protein extraction and separation has been reported using the ionic liquid (IL)-based aqueous two-phase system (ATPS). Although ILs have many benefits, there was a problem of unfavorable physical properties such as high viscosity, or high-cost production of ILs. Recently protic ionic liquids (PILs) are emerging as an alternative to conventional ionic liquids. PILs are easily produced through a reaction between Brønsted acid and base at a relatively low cost. In this study, PILs (pyrrolidinium formate [Pyrr][HCOO] and propionate [Pyrr][C 2 H 5 COO] and inorganic salts (K 3 PO 4 , K 2 HPO 4 ) as phase-forming constituents of ATPS, were used for the protein extraction. From the binodal experiments, the phase separation abilities are [Pyrr][HCOO] > [Pyrr][C 2 H 5 COO] and K 3 PO 4 > K 2 HPO 4 . Because proteins precipitated at the interface when using K 3 PO 4 , ATPS composed of [Pyrr][HCOO] and K 2 HPO 4 was selected for the protein extraction. The proteins were successfully extracted to PIL-rich phase. The distribution ratio of each protein was in the order hemoglobin < cytochrome C <  chymotrypsin < albumin. The partition behavior of proteins in PIL-based ATPS was complex and influenced by combined effect of hydrophobic, electrostatic interactions and salting out. FT-IR spectroscopic characterization indicated that secondary structures of hemoglobin and cytochrome C were retained in PIL-rich phase after