In this study an aqueous two-phase system (ATPS) composed of polyethylene glycol (PEG) and potassium phosphate was tested for the purification of lipase from Yarrowia lipolytica IMUFRJ 50682. Ultrafiltration and precipitation with acetone and kaolin were also used as traditional comparison methods Ultrafiltration was a good method with a purification factor of 6.55, but protease was also purified in this extract. For the precipitation with acetone and kaolin lower values of lipase and protease activity were found in relation to the original crude enzyme extract. Under the best conditions of ATPS (pH 6 and 4 degrees C), the purification fold was greater than 40 and selectivity was almost 500. Lipase was recovered in the salty phase which makes it easier to purify it. The optimum pH and temperature ranges for purified lipase with this system was 6-7 and 35-40 degrees C, respectively. Lipase thermostability was increased in relation to crude extract after the purification with the PEG/phosphate buffer system for temperatures lower than 50 degrees C. All enzyme extracts showed good stability to a wide pH range. Y. lipolytca lipase was successfully purified by using ATPS in a single downstream processing step and presented good process characteristics after this treatment. (C) 2016 Elsevier Ltd. All rights reserved.