Protein partitioning in thermoseparating systems of a charged hydrophobically modified ethylene oxide polymer

• Published in: Journal of Chromatography A Vol. 983; no. 1; pp. 133 - 144
• Main Authors: Jönsson, Malin, Johansson, Hans-Olof
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 03.01.2003; Elsevier
• Subjects: aqueous two-phase systems; Biologi; Biological and medical sciences; Biological Sciences; Biotechnology; Ethylene Oxide - chemistry; Fundamental and applied biological sciences. Psychology; Methods. Procedures. Technologies; mixed micelles; Natural Sciences; Naturvetenskap; Others; Poly(ethylene oxide); Polymers - chemistry; protein partitioning; Proteins; Proteins - isolation & purification; Sodium Dodecyl Sulfate; temperature effects; thermoseparation modelling; Various methods and equipments; Mixed micelles; Proteins; Protein partitioning; Temperature effects; Aqueous two-phase systems; Thermoseparation modelling; Poly(ethylene oxide); Cationic surfactant; Temperature effect; Mixed micelle; Partition coefficient; Modeling; Separation method; pH; Ungulata; Amine copolymer; Lysozyme; Purification; Bovine; Phase composition; Enzyme; Serum albumin; Ethylene oxide copolymer; Serum protein; Vertebrata; Biphasic system; Mammalia; Salt effect; Glycosidases; Animal; Hydrolases; Artiodactyla; Aqueous solution; O-Glycosidases
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(02)01695-3

The phase behavior of a thermoseparating cationic hydrophobically modified ethylene oxide polymer (HM-EO) containing tertiary amines has been investigated at different pH, salt and sodium dodecyl sulfate (SDS) concentrations, in order to find a water/HM-EO two-phase system suitable for protein partitioning. The used polymer forms micellar aggregates that can be charged. By changing pH and SDS concentrations the netcharge of the SDS/HM-EO aggregate can be shifted from positive to negative. Bovine serum albumin (BSA) and lysozyme were partitioned in the thermoseparated two-phase systems of the cationic polymer at different pH, salt and SDS concentrations. The dominant attractive interactions between the polymer aggregates and the studied proteins were shown to be of electrostatic (Coulomb) nature rather than hydrophobic interaction. At low ionic strength the positively charged polymeric aggregates attracted negatively charged BSA and repelled positively charged lysozyme. Upon addition of SDS the negatively charged aggregates attracted lysozyme and repelled BSA. Thus, it was possible to direct proteins with different charges to the polymeric phase and redirect them to a polymer-depleted phase by changing the netcharge of the polymeric aggregates. The effect of different salts on the partitioning of BSA in a system of slightly positively charged HM-EO was studied. NaCl and KBr have a significant effect on driving the BSA to the polymer-depleted phase, whereas KF and K 2SO 4 have a smaller effect on the partitioning. The cloud point temperature of the charged polymer decreased upon addition of SDS near the isoelectric molar ratio of SDS to polymer and also upon salt addition. In the latter case the decrease was smaller than expected from model calculations based on Flory–Huggins theory, which were performed for a charged thermoseparating polymer at different charges and salt concentrations.