Abstract
A series of new self-buffering ionic liquids (ILs) based on Good's buffers (GBs) anions and the tetrabutylphosphonium cation ([P4444]+) was here synthesized and characterized. The self-buffering behaviour of the GB-ILs was confirmed by measuring their protonation constants by potentiometry. Further, their ability to form aqueous biphasic systems with the biodegradable potassium citrate salt was evaluated, and further investigated for the extraction of proteins, using bovine serum albumin (BSA) as a model protein. If these ionic structures display self-buffering characteristics as well as a low toxicity towards the luminescent bacteria Vibrio fischeri, they were additionally found to be highly effective in the formation of ABS and in the extraction of BSA - extraction efficiencies of 100% to the IL-rich phase obtained in a single-step. The BSA secondary structure in the aqueous IL-rich solutions was evaluated through infrared spectroscopic studies revealing the protein-friendly nature of the synthesized ILs. Dynamic light scattering (DLS), "COnductor-like Screening MOdel for Real Solvents" (COSMO-RS), and molecular docking studies were finally carried out to better understand the main driving forces of the extraction process. The results suggest that van der Waals and hydrogen-bonding interactions are important driving forces of the protein migration towards the GB-IL-rich phase, while the molecular docking investigations demonstrated a stabilizing effect of the studied ILs over the protein.
Highlights
Aqueous biphasic systems (ABS) are more benign alternatives to replace the conventional liquid–liquid extractions using organic volatile solvents aiming the efficient separation of biomolecules, in which proteins and enzymes are included (Asenjo and Andrews, 2011; Benavides and Rito-Palomares, 2008)
The pH-profiles of these Good’s buffers (GBs)-ionic liquids (ILs) were measured in aqueous solution (Figure 2a) to identify their buffering action, through which is possible to show their buffering-like region described by the moderate slope length appearing before the inflection point at high pH
We can see that the buffer capacities of GB-ILs are relatively high and offer a wide range of pH values
Summary
Aqueous biphasic systems (ABS) are more benign alternatives to replace the conventional liquid–liquid extractions using organic volatile solvents aiming the efficient separation of biomolecules, in which proteins and enzymes are included (Asenjo and Andrews, 2011; Benavides and Rito-Palomares, 2008). ABS are claimed as greener extraction processes, minimizing the consumption of volatile organic solvents which could be harmful to the environment and human resources, and for proteins since they could cause their denaturation (Krishna et al, 2002) Several techniques such as electrophoresis, membrane separation, gel filtration, and affinity chromatography, were adopted in protein separation; they are some of the most expensive processes and are associated to several limitations, namely concerning their scale-up (Pei et al, 2009). Traditional ABS consist of aqueous solutions of two polymers (e.g. polyethylene glycol and dextran) or a polymer and a salt They have been established as an economical method making part of downstream processes exhibiting low-energy consumption, high performance, high biocompatibility and suitability for large scale production. The ABS processes have been extensively studied for the separation and purification of different biological products
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