Halogenated arylamines are key building blocks in the production of fine chemicals and pharmaceuticals. As a result, there is growing interest in the use of enzymes to enable improved regioselectivity and eco-friendliness in the synthesis of this class of compounds. Currently, there are several research programmes on the development of halogenase enzymes as biocatalysts for this purpose, but progress in this area is hampered by a lack of suitable high-throughput screening methods.Here, the use of a peroxidase-mediated assay based on the coupling of oxidised 4-methyl-catechol to the arylamines is reported. The coupling reaction results in brightly coloured products from which UV-Vis spectrometry readily differentiates between different halogenated products and unhalogenated starting materials, enabling the rapid identification of successful halogenation reactions. The colour generated is also observable by eye and the assay is compatible with high-throughput UV-Vis analysis in a microtitre plate format.Furthermore, the assay is also able to provide accurate quantification down to 50�M concentrations by using a ratiometric method of analysis; and can be implemented as an end point assay with flavin-dependent halogenases such as RebH, enabling in situscreening.?

Enzyme-compatible, high-throughput, and quantitative spectrophotometric assays for arylamine halogenations

248th National Meeting (2014)

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