Journal of Scientific & Industrial Research Vol 74, June 2015, pp. 348-353 Extraction of bovine serum albumin using aqueous two-phase poly (ethylene glycol) – poly (acrylic acid) system S Settu, P Velmurugan, R R Jonnalagadda * and B U Nair Chemical Laboratory, Council of Scientific and Industrial Research-Central Leather Research Institute, Adyar, Chennai - 600 00, India Received 6 August 2014; revised 2 February 2015; accepted 12 April 2015 The partitioning behavior of bovine serum albumin in a new aqueous two-phase system (ATPS) based on low cost commercial polymers like poly (ethy1ene glycol) (PEG) and poly (acrylic acid) (PAA) has been studied. In this ATPS, PAA and PEG are enriched in the bottom and upper phase, respectively. The influence of molecular weight (MW), tie line length, pH, temperatures and NaCl concentration on the partition coefficient of protein has been studied. The protein partitioning decreases on increasing MW of PEG and temperature, whereas increases with an increasing pH and NaCl concentration. This may be attributed by strong electrostatic interactions between the proteins and polymer as well as excluded volume effects. Protein partitioning is better for PEG4000 at 0 o C, pH 8.0 with 1 M NaCl with a yield 88.8%. The present study indicates that PEG-PAA ATPS may be considered as an interesting alternative for protein purification from the biological suspensions. Keywords: Aqueous Two Phase; Liquid-Liquid Extraction; Protein Separation; Poly (ethylene glycol); Poly (acrylic acid); Bovine serum albumin Introduction Aqueous two-phase systems (ATPS) are a powerful method in biochemical research for the separation and purification of macromolecules, cells, and cell fragments. Conventional methods used for protein purification are usually expensive and difficult to scale up. For this reason, a need exists for an efficient, effective and economic large-scale bio-separation technique to achieve high purity and high recovery, while maintaining the biological activity of the protein. Hence, there is an ongoing interest in biotechnology for the development of new separation and purification methods that are both economically viable and gentle enough to preserve biological activity of proteins. ATPS is one of the novel techniques to provide secured separation and purification of bio-molecules 1-3 . The most commonly used and investigated ATPS are composed of either PEG–inorganic salts or PEG – dextran. It also has the disadvantage of low solubility for amphiphilic proteins and high salt concentration results in waste disposal problem 7 . To overcome these problems, polymer-polymer systems are being used for protein extraction. High cost of dextran leads to continuous interest for developing novel polymers 4-7 . The PEG-PAA aqueous two-phase polymer-polymer system has been developed and well characterized by our group 8 . The research presented here will raise the potential practical application of a previously well-characterized system. An attempt has been made to study the partitioning behavior of model protein bovine serum albumin in order to evaluate the capability of the novel aqueous two-phase poly (ethylene glycol) (PEG) – poly (acrylic acid) system for protein separation. Materials and methods Materials Polyethylene glycol with molecular weights of 4000, 6000 and 10000 (Da) was obtained from Merck-Schuchardt (Munich, Germany). Bovine serum albumin (BSA) was purchased from Sigma (St. Louis, MO, USA). Poly (acrylic acid, sodium salt) of average molecular weight of 100 was procured from Aldrich chemicals company, USA. For the present work the polymers were used without further purification. Milli-Q water was used throughout the experiments. Circular dichroism spectrum of BSA Dichroic spectroscopy measurements were conducted on a JASCO J-815 CD spectrophotometer (JASCO Inc.) for pure and extracted BSA. For CD ___________  Author for correspondence E-mail: jrrao@clri.res.in