title: Preparation of labeled human drug metabolites and drug-drug interaction-probes with fungal peroxygenases.
abstract: Enzymatic conversion of a drug can be an efficient alternative for the preparation of a complex metabolite compared with a multi-step chemical synthesis approach. Limitations exist for chemical methods for direct oxygen incorporation into organic molecules often suffering from low yields and unspecific oxidation and also for alternative whole-cell biotransformation processes, which require specific fermentation know-how. Stable oxygen-transferring biocatalysts such as unspecific peroxygenases (UPOs) could be an alternative for the synthesis of human drug metabolites and related stable isotope-labeled analogues. This work shows that UPOs can be used in combination with hydrogen/deuterium exchange for an efficient one-step process for the preparation of 4'-OH-diclofenac-d6. The scope of the reaction was investigated by screening of different peroxygenase subtypes for the transformation of selected deuterium-labeled substrates such as phenacetin-d3 or lidocaine-d3. Experiments with diclofenac-d7 revealed that the deuterium-labeling does not affect the kinetic parameters. By using the latter substrate and H2 (18) O2 as cosubstrate, it was possible to prepare a doubly isotope-labeled metabolite (4'-(18) OH-diclofenac-d6). UPOs offer certain practical advantages compared with P450 enzyme systems in terms of stability and ease of handling. Given these advantages, future work will expand the existing 'monooxygenation toolbox' of different fungal peroxygenases that mimic P450 in vitro reactions.
authors.affiliation: Department of Biological and Environmental Sciences, TU Dresden - International Institute Zittau, Markt 23, 02763, Zittau, Germany.
authors.forename: Marzena
authors.initials: M
authors.lastname: Poraj-Kobielska
authors.forename: Jens
authors.initials: J
authors.lastname: Atzrodt
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Holla
authors.forename: Martin
authors.initials: M
authors.lastname: Sandvoss
authors.forename: Glenn
authors.initials: G
authors.lastname: Gröbe
authors.forename: Katrin
authors.initials: K
authors.lastname: Scheibner
authors.forename: Martin
authors.initials: M
authors.lastname: Hofrichter
mesh_terms: D000363:Agaricales / Q000201:enzymology*; D041961:Deuterium Exchange Measurement; D004347:Drug Interactions; D006801:Humans; D006900:Hydroxylation; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D015335:Molecular Probes / Q000378:metabolism*; D004364:Pharmaceutical Preparations / Q000737:chemistry / Q000378:metabolism*
chemical_list: D015335:Molecular Probes; D004364:Pharmaceutical Preparations; D006899:Mixed Function Oxygenases; C022333:peroxygenase
keywords: 4'OH-diclofenac; EC 1.11.2.1; H/D exchange; Isotopic labeled synthesis; deuterium; human drug metabolites; lidocaine; paracetamol; phenacetin; unspecific/aromatic peroxygenase