PEG–salt aqueous two-phase systems: an attractive and versatile liquid–liquid extraction technology for the downstream processing of proteins and enzymes

Abstract

Nowadays, there is an increasing demand to establish new feasible, efficient downstream processing (DSP) techniques in biotechnology and related fields. Although several conventional DSP technologies have been widely employed, they are usually expensive and time-consuming and often provide only low recovery yields. Hence, the DSP is one major bottleneck for the commercialization of biological products. In this context, polyethylene glycol (PEG)-salt aqueous two-phase systems (ATPS) represent a promising, efficient liquid-liquid extraction technology for the DSP of various biomolecules, such as proteins and enzymes. Furthermore, ATPS can overcome the limitations of traditional DSP techniques and have gained importance for applications in several fields of biotechnology due to versatile advantages over conventional DSP methods, such as biocompatibility, technical simplicity, and easy scale-up potential. In the present review, various practical applications of PEG-salt ATPS are presented to highlight their feasibility to operate as an attractive and versatile liquid-liquid extraction technology for the DSP of proteins and enzymes, thus facilitating the approach of new researchers to this technique. Thereby, single- and multi-stage extraction, several process integration methods, as well as large-scale extraction and purification of proteins regarding technical aspects, scale-up, recycling of process chemicals, and economic aspects are discussed.

A detailed overview of xylanases: an emerging biomolecule for current and future prospective

“…The 60-70% of total processing cost in enzyme downstream process comes from the purification step (Loureiro et al 2017;Bhardwaj et al 2019). Therefore, several scientists suggested a single step liquid-liquid fractionation technique, i.e., aqueous two-phase system (Naganagouda and Mulimani 2008;Yasinok et al 2010;Glyk et al 2015). Garai and Kumar (2013) purified alkaline xylanase from Aspergillus candidus using aqueous two-phase system (ATPS) composed of PEG 4000/NaH 2 PO 4 system.…”

Programmable Hydrogel Ionic Circuits for Biologically Matched Electronic Interfaces

“…The hydrogel ionic circuits were developed based on ATPS . PEG is commonly used in ATPS, which can phase separate with various salts .…”

Evaluation of Driving Forces for Protein Partition in PEG-Salt Aqueous Two- Phase Systems and Optimization by Design of Experiments

“…Increasing KPP (X 3 ) and NaCl (X 4 ) concentration had positive or negative effects, and rising PEG (X 2 ) concentration as well as PEG MW (X 1 ) had negative or positive effects on the partitioning or recovery yield of BSA/OVA (Figures 3A/S6A and 4A/S7A), respectively, while increasing NaCit (X 3 ), NaCl (X 4 ) and PEG concentration ( Since BSA/OVA are predominantly partitioned into the salt-rich bottom phase (K<1) as well as LYZ/α-CT into the PEG-rich top phase (K>1), the distribution of BSA/OVA into the bottom phase or LYZ/α-CT into the top phase is more pronounced at lower or higher values of K, respectively. Thereby, the partitioning behavior and recovery yields of BSA/OVA were influenced by negative or positive interaction effects of factors PEG MW × NaCl (X 1 × X 4 ) and PEG × NaCl (X 2 × X 4 Overall, the effects of significant factors/interactions regarding a decreasing influence on the protein partitioning and recovery were ranked in the following order: KPP/NaCit ( (Tables S4-S7; Figures 3, 4, S6 and S7). As a conclusion from the 2 5-1 fFD, the effects of KPP/NaCit, NaCl and PEG concentration were confirmed as the significant factors and therefore selected for further optimization experiments in order to improve the response variables of each investigated model protein.…”

“…ATPS provide several advantages over conventional downstream processing techniques for separation, recovery and purification of biomolecules, such as biocompatibility, technical simplicity, high capacity, as well as easy scalability. Considering these advantages, ATPS have been used since the mid-1950s as a mild liquid-liquid extraction and bio separation technology of wide applicability in biotechnology, biochemistry, and cell biology, such as large-scale separation, partial purification or primary recovery of various biological products like proteins, enzymes and nucleic acids [1][2][3][4]. Among all systems suitable for bio separation, PEG-salt ATPS have been used predominantly concerning the low-cost of the phase-forming constituents, low viscosity and short separation time [1,5].…”