Aqueous two-phase systems in the presence of denaturants (urea or guanidine hydrochloride) have been used for the initial recovery steps during protein refolding process. To investigate phase behavior of such systems, liquid-liquid equilibrium data for a poly (ethylene glycol) (PEG) 4000 + sodium sulfate + urea/guanidine hydrochloride (5% and 10% mass) aqueous two phase system was investigated at 25 degrees C and pH values of 5.3 and 9.2. The calibration plots for refractive index and conductivity were prepared to obtain the compositions of the top and bottom phase. It was observed that the heterogeneous region decreases with the increase in the denaturants concentration. The effect of pH, denaturants concentration, tie line length, and slope of tie line on the partition behavior of denaturants was examined. In this study, the modified UNIQUAC-FV and extended UNIQUAC models are used to calculate the phase equilibria of mention aqueous two phase systems. The values of average AAD% (partition coefficients) for modified UNIQUAC-FV and extended UNIQUAC are 3.48 and 13.80, respectively. (C) 2016 Elsevier B.V. All rights reserved.