Design of green DES-based aqueous two phase systems for lipolytic enzymes extraction

• A biocompatible process using reline DES was developed for lipase extraction. • Phase separation with potassium organic salts and reline in Tergitol 15-S9 was done. • Superior biocompatibility of reline with three commercial lipases was demonstrated. • About 90% extraction efficiency was attained after reline-based ABS was performed. This study pioneers a novel green and cost-effective separation process using reline deep eutectic solvent (DES) for extracting Thermomyces lanuginosus lipase ( Tl L) from aqueous solutions. For the first time, it has been shown that various potassium organic salts combined with reline DES can induce phase separation in aqueous solutions of the biodegradable non-ionic surfactant Tergitol 15-S9, presenting a groundbreaking alternative to the banned surfactant Triton X-100. This work uniquely applies three different equations to describe the experimental solubility curves. The selection of reline DES is particularly innovative as it avoids significant deactivation effects with three commercial lipases: Candida antarctica lipases A and B, and Tl L, with the latter undergoing a detailed study. This includes an unprecedented analysis of its thermodeactivation kinetics in reline DES aqueous solutions. Additionally, a novel investigation of immiscibility regions at different temperatures, involving comprehensive Tie-Line characterizations (slope and length), preceded the evaluation of lipase extraction. Remarkably, the study demonstrates that optimizing the feed composition to contain higher water concentrations can achieve exceptionally high extraction efficiencies (approximately 90%), suggesting a transformative potential for future applications in real aqueous cultures.