Aqueous two‐phase system based on natural quaternary ammonium compounds for the extraction of proteins

Aqueous two‐phase systems, based on the use of natural quaternary ammonium compounds, were developed to establish a benign biotechnological route for efficient protein separation. In this study, aqueous two‐phase systems of two natural resources betaine and choline with polyethyleneglycol (PEG400/600) or inorganic salts (K₂HPO₄/K₃PO₄) were formed. It was shown that in the K₂HPO₄‐containing aqueous two‐phase system, hydrophobic interactions were an important driving force of protein partitioning, while protein size played a vital role in aqueous two‐phase systems that contained polyethylene glycol. An extraction efficiency of more than 90% for bovine serum albumin in the betaine/K₂HPO₄ aqueous two‐phase system can be obtained, and this betaine‐based aqueous two‐phase system provided a gentle and stable environment for the protein. In addition, after investigation of the cluster phenomenon in the betaine/K₂HPO₄ aqueous two‐phase systems, it was suggested that this phenomenon also played a significant role for protein extraction in this system. The development of aqueous two‐phase systems based on natural quaternary ammonium compounds not only provided an effective and greener method of aqueous two‐phase system to meet the requirements of green chemistry but also may help to solve the mystery of the compartmentalization of biomolecules in cells.