Regioselective synthesis of 3-hydroxyorthanilic acid and its biotransformation into a novel phenoxazinone dye by use of laccase

A natural phenoxazinone derivative, cinnabarinic acid (2, CA, R = CO2H), could be obtained in vitro with the aid of purified laccase from the fungi Pycnoporus cinnabarinus by oxidative dimerization of 3-hydroxyanthranilic acid (1, 3-HAA, R = CO2H). With the aim of gaining access to a new class of water-soluble chromophores and potential bioactive molecules, the regioselective sulfonation of 2-hydroxyanihne was investigated. Straightforward insertion of a sulfonate group into a carbon-metal bond provided an efficient and selective process for the synthesis of 3-hydroxyorthanilic acid (3, 3-HOA, R = SO3H). This sulfonated compound was then subjected to laccase oxidation in order to mimic the cinnabarinic acid synthesis. A practical HPLC method to study the oxidized products was developed, and the major product of biotransformation - namely 2-aniino-3-oxo-3H-phenoxazin-1,9-disulfonic acid (4, LAO, R = SO3H) - was isolated and identified as the sulfonate analogue of cinnabarinic acid. ((c) Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008).