Publication: Protein extraction using an aqueous quaternary two-phase system: the role of hydrogen bond donor
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Date
2023-08-01
Authors
Saravanakumara A/L Marappan
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Abstract
Liquid – liquid extraction such as aqueous quaternary two-phase system (AQTPS) has been a promising method in the biotechnology field for biomolecules extraction. The binary mixture of PEG/salt is the most common aqueous two-phase system (ATPS). However, naturally derived binary mixtures like deep eutectic solvents (DES) based AQTPS have attracted more attention of researchers for biomolecule extraction. However, the ability of eutectic mixture-based AQTPS to fractionate protein macromolecules remains not well understood. This is due to the variety of hydrogen bond donor (HBD) used to synthesize DESs. Hence, the role of HBDs in eutectic mixture was investigated in relation to the phase formation ability and extraction efficiency of protein. In this study the AQTPSs were composed of potassium salt and eutectic mixture. Six types of deep eutectic mixture were prepared, which are ChCl-urea, ChCl-glycerol, ChCl-sorbitol, ChCl-fructose, ChCl-glucose and ChCl-1,3-propanediol. All of the eutectic mixture synthesized are able to form two phases when added to potassium salt solution. It was found out the highest extraction efficiency of protein was achieved by using the ChCl-urea AQTPS followed by ChCl-1,3-propanediol AQTPS. On top of that, in terms of phase forming ability ChCl -urea and ChCl -1,3-propanediol resulted in curve closest to the axis of origin establishing its stronger phase forming ability in AQTPS. In addition to that, it was observed that polyol-based DES namely ChCl -sorbitol has tremendous effect on the resulting biphasic region. This was due to the reducing volume of DES rich top phase when additional volume of salt solution was introduced to the AQTPS. Eventually, the extraction performance and phase forming ability of DESs were found out to be strongly related to the hydrophobicity/hydrophilicity of the HBDs used. Besides, the DESs which could possibly act as a phase former, or modifier were screened based on the hydrophobicity of HBDs. Thus, it confirmed that varying the HBDs gives significant impact on the eutectic mixtures based AQTPS. Regardless, findings from this study proved that eutectic mixtures based two phase system has potential for the extraction of protein owing to its excellent extraction performance.