Continuous oxyfunctionalizations catalyzed by unspecific peroxygenase

Unspecific peroxygenase (UPO) has been shown to be a promising biocatalyst for oxyfunctionalization of a broad range of substrates with hydrogen peroxide (H 2 O 2 ) as the cosubstrate. Unspecific peroxygenase (UPO) has been shown to be a promising biocatalyst for oxyfunctionalization of a broad range of substrates with hydrogen peroxide (H 2 O 2 ) as the cosubstrate. In this study, we used the UPO mutant PaDa-I from Agrocybe aegerita ( Aae UPO) for the enantiopure synthesis of ( R )-1-phenylethanol from ethylbenzene and transferred it into continuously operated reactors. While the free enzyme was applied in an enzyme membrane reactor (EMR), the enzyme was immobilized on the ECR8315F carrier material and used in a packed-bed reactor (PBR). Both systems were fully characterized (residence time distributions, RTD) and a standard test reaction, the oxidation of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), was carried out in continuous operation. Throughout, we found 1.5- and 1.2-times higher experimental RTDs than theoretical RTDs for the EMR and PBR, respectively. The EMR showed simplicity of handling, but it could not withstand the demands of a robust continuous synthesis. Therefore, the focus was on the PBR. We optimized the reaction conditions and produced 272 mg of ABTS˙ + -radical in 6.9 days and achieved a space–time-yield (STY) of 0.68 g (L h) −1 and a productivity of 0.16 mg (L h) −1 . Eventually, we demonstrated the oxyfunctionalization of ethylbenzene to ( R )-1-phenylethanol in continuous operation for the first time in the literature with a STY of 0.97 g (L h) −1 and a productivity of 0.25 mg (L h) −1 . Hydrogen peroxide was stoichiometrically converted and the system proceeded for 6.2 days and neither the overoxidation byproduct acetophenone, nor the ( S )-enantiomer was detected.