Fundamental studies of biomolecule partitioning in aqueous two‐phase systems

Abstract

Phase diagram data at 4 degrees C was determined for the aqueous two-phase systems composed of polyethylene glycol, dextran, and water. The Flory-Huggins theory of polymer thermodynamics was used to correlate partitioning of biomolecules in these aqueous two-phase systems resulting in a simple linear relationship between the natural logarithm of the partition coefficient and the concentration of polymers in the two phases. This relationship was verified by partitioning a series of dipeptides which differ from one another by the addition of a CH(2) group on the c-terminal amino acid residue and by utilizing a set of low-molecular-weight proteins. The slope of the line could be expressed in terms of the interactions of the biomolecule with the phase forming polymers and water. The main result for the dipeptides was that knowledge of the partition coefficient in any of the PEG/dextran/water systems, regardless of polymer molecular weight, enabled prediction of the coefficient in all of the systems. The dipeptides were also used for determination of the Gibbs free energy of transfer of a CH(2) group between the phases. This quantity was correlated with polymer concentration, thus establishing a hydrophobicity profile for the PEG/ dextran/water systems. The methodology for predicting dipeptide partition coefficients was extended to proteins, where it was found that low-molecular-weight proteins gave a linear relationship with the tie line compositions of a phase diagram.

Contribution of protein charge to partitioning in aqueous two-phase systems

“…Where ⌬ is proportional to the tie-line length (TLL) (Diamond and Hsu, 1989;Hartounian et al, 1994;King et al, 1988), defined as…”

Application of integral‐equation theory to aqueous two‐phase partitioning systems

“…Also, Diamond and Hsu (1989) correlated an extensive set of dipeptide and low-molecular-weight protein partition-coefficient data using a simple, linearized form of Flory-Huggins theory.…”

Partitioning of cerrena unicolor laccase activity in an aqueous two-phase system

“…One of the methods to overcome this problem is using aqueous two-phase systems (ATPS) for protein purification, because of their ability to extract macromolecules without deactivating them, the ease of scale-up and usually high activity yields and partitioning coefficients (Diamond and Hsu, 1989;Silvério et al, 2012). Aqueous two-phase systems consist of two non-miscible aqueous phases, which are formed when two mutually immiscible polymers, or a polymer with a salt, an alcohol and a salt, an ionic liquid and a salt, or two surfactants solutions are introduced into the system (Salabat et al, 2010).…”