Environmentally Benign and Recyclable Aqueous Two-Phase System Composed of Distillable CO₂-Based Alkyl Carbamate Ionic Liquids

Ionic liquid-based aqueous two-phase systems (IL-ATPSs) have been studied comprehensively as a potential alternative method for protein purification. One of the distinct features of IL-ATPS is that the polarity of the IL can be adjusted by different combinations of cation and anion, providing flexibility in governing the partition of proteins between phases. However, the conventional IL-ATPS usually has poor environmental footprint and low recyclability, thereby hampering the systems for practical use in protein separation. To address these shortcomings, here we explored the distillable CO₂-based alkyl carbamate ILs as phase-forming components. N,N-dimethylammonium N′,N′-dimethylcarbamate (DIMCARB) was able to form ATPS with polypropylene glycol (PPG). The liquid-liquid equilibrium data of ATPSs composed of PPG with different molecular mass and DIMCARB were determined. As both phase-forming components are thermosensitive, the effect of temperature on the phase diagram of the PPG 400 + DIMCARB system was also evaluated. The partition behavior of model proteins (i.e., bovine serum albumin and lysozyme) in the PPG + DIMCARB systems were assessed. Both model proteins tend to migrate to the more hydrophilic IL-rich phase, when a higher temperature or a higher molecular mass of PPG was used. The DIMCARB was recovered via rotary evaporation, while the PPG 400 was recovered from a secondary two-phase system thermally induced from the primary ATPS. The recovered components were subsequently used to prepare a new batch of ATPS. The phase composition of the new system as well as the partition coefficients of both model proteins in the recycled system were maintained, thereby proving the feasibility of reusing both phase-forming components. Overall, the recyclable and biodegradable phase-forming components used in the newly developed PPG + DIMCARB system make the ATPS a sustainable and eco-friendly system for protein purification.