Mechanistic Insights into the Direct Enzymatic Conversion of Oleuropein to Oleacein from Olive Leaves: Natural Deep Eutectic Solvents as a Tunable Reaction Environment

Traditional methods for obtaining oleacein from olive oil or fruits are often inefficient, tedious, labor-intensive, time-consuming, and potentially hazardous. This study first demonstrated that the natural deep eutectic solvent (NADES) composed of choline chloride, glycerol, and water enabled highly efficient and selective conversion of oleuropein into oleacein within olive leaves. Mechanistic analysis revealed an enzymatic pathway in which the glycerol-based NADES system promoted endogenous enzyme activities (particularly β-glucosidase), thereby facilitating the targeted conversion. The optimal choline chloride/glycerol/water composition (15:29:56, wt %) exhibited a pH of 5.4, which matched the optimal pH of both olive β-glucosidase and the leaf apoplast, implying a cytosol-like microenvironment and yielding 51.46 mg/g DW of oleacein under ultrasonic assistance. Collectively, these findings extend the role of NADES as a versatile reaction environment for the targeted conversion of secondary metabolites and provide a new strategy for the large-scale acquisition of oleacein and the high-value utilization of olive leaf byproducts.