Purification and recovery of laccase produced by submerged cultures of Trametes versicolor by three-phase partitioning as a simple and highly efficient technique

In this work, three-phase partitioning (TPP) was used for the purification of laccase from liquid cultures of Trametes versicolor. For determining the optimal conditions of TPP process, parameters such as initial pH (6.5, 7.0, 7.5, 8.0), ammonium sulphate saturation (20%-80%) and the water phase to tert-butanol ratio (1:0.5, 1:1, 1:2), were analyzed. The best conditions with 73% recovery and 24-fold purification was obtained with the use of 50% saturation with ammonium sulphate, water phase to tert-butanol ratio of 1:1 and initial pH 7.0. The molecular mass of the purified laccase secreted by analyzed strain T. versicolor was found for ≈ 66 kDa. The results showed that TPP is an efficient method for the fractionation and purification of laccase obtained from liquid cultures of T. versicolor and it allows for obtaining the relatively pure enzyme without the use of time-consuming and costly chromatographic methods.