Terholsen_2022_Chembiochem_23_e202200254

Olive mill wastewater (OMWW) is produced annually during olive oil extraction and contains most of the health-promoting 3-hydroxytyrosol of the olive fruit. To facilitate its recovery, enzymatic transesterification of hydroxytyrosol (HT) was directly performed in an aqueous system in the presence of ethyl acetate, yielding a 3-hydroxytyrosol acetate rich extract. For this, the promiscuous acyltransferase from Pyrobaculum calidifontis VA1 (PestE) was engineered by rational design. The best mutant for the acetylation of hydroxytyrosol PestE_I208A_L209F_N288A was immobilized on EziG2 beads, resulting in hydroxytyrosol conversions between 82 and 89% in one hour, for at least ten reaction cycles in a buffered hydroxytyrosol solution. Due to inhibition by other phenols in OMWW the conversions of hydroxytyrosol from this source were between 51 and 62%. In a preparative scale reaction, 13.8 mg (57%) of 3-hydroxytyrosol acetate was extracted from 60 mL OMWW.