Purification of plant-esterase in PEG1000/NaH2PO4 aqueous two-phase system by a two-step extraction

Purification of plant-esterase from flour in an aqueous two-phase system (ATPS) was investigated. The effects of various process parameters such as the type of aqueous two-phase systems, the phase-forming salt, the molecular weight and concentration of PEG, the system pH, and the types and concentrations of neutral salts on partitioning of plant-esterase were evaluated. Optimized conditions for the purification of plant-esterase were found in polymer–salt systems, with especially promising results in the PEG1000/NaH2PO4 system. Using 27.0% PEG1000/13.0% NaH2PO4 (w/w, pH 5.0), and 27.0% PEG1000/13.0% NaH2PO4/6.0% (NH4)2SO4 (w/w, pH 5.0), plant-esterase was purified by a two-step extraction. Compared to the results obtained with the conventional salting-out method, this method had a comparable yield (83.16% versus the original yield of 80%), but produced plant-esterase that was 4.8 times as pure (18.46-fold). Integrating dialysis into the aqueous two-phase extraction removed (NH4)2SO4 from the purified plant-esterase. Finally, plant-esterase was freeze-dried to convert the product to powder. This work offers a simple and more efficient process to purify and concentrate plant-esterase. Plant-esterase is used in applications such as organophosphorus compounds (OPs) detection and since our method makes this enzyme easier to isolate, it will enhance researchers’ ability to explore these applications.