Synthesis of oxyfunctionalized NSAID metabolites by microbial biocatalysts

The synthesis of valuable metabolites and degradation intermediates of drugs, like non-steroidal anti-inflammatory drugs (NSAIDs), are substantially for toxicological and environmental studies, but efficient synthesis strategies and the metabolite availability are still challenging aspects. To overcome these bottlenecks filamentous fungi as microbial biocatalysts were applied. Different NSAIDs like diclofenac, ibuprofen, naproxen and mefenamic acid could be oxyfunctionalized to produce human metabolites in isolated yields of up to 99% using 1 g L-1 of substrate. Thereby the biotransformations using Beauveria bassiana, Clitocybe nebularis or Mucor hiemalis surpass previous reported chemical, microbial and P450-based routes in terms of efficiency. In addition to different hydroxylated compounds of diclofenac, a novel metabolite, 3,4-dihydroxydiclofenac, has been catalyzed by B. bassiana and the responsible P450s were identified by proteome analysis. The applied filamentous fungi present an interesting alternative, microbial biocatalysts platform for the production of valuable oxyfunctionalized drug metabolites.\n\nImportanceThe occurrence of pharmaceutically active compounds, such as diclofenac and its metabolites, in the environment, in particular in aquatic systems, is of increasing concern because of the increased application of drugs. Standards of putative metabolites are therefore necessary for environmental studies. Moreover, pharmaceutical research and development requires assessment of the bioavailability, toxicity and metabolic fate of potential new drugs to ensure its safety for users and the environment. Since most of the reactions in the early pharmacokinetics of drugs are oxyfunctionalizations catalysed by P450s, oxyfunctionalized metabolites are of major interest. However, to assess these metabolites chemical synthesis often suffer from multistep reactions, toxic substances, polluting conditions and achieve only low regioselectivity. Biocatalysis can contribute to this by using microbial cell factories. The significance of our research is to complement or even exceed synthetic methods for the production of oxyfunctionalized drug metabolites.