Enantioselective toluene dioxygenase catalysed di- and tri-hydroxylation of monosubstituted benzenes

Asymmetric cis-dihydroxylation to yield diols 2A–2G and sequential benzylic monohydroxylation–cis-dihydroxylation to yield triols 4A–4G (trihydroxylation), occurred during biotransformation of a series of monosubstituted alkylbenzene substrates 1A–1G using toluene dioxygenase, a biocatalyst present in Pseudomonas putida UV4. Dioxygenase-catalysed cis-dihydroxylation of the R and S benzylic alcohol enantiomers 3B–3D, 3B′–3D′ gave the corresponding enantiopure triols 4B–4D, 4B′–4D′. Biotransformation of substrates 1J–1L yielded cis-diols 2J–2L and a minor triol metabolite 4A. Benzylic alcohols 3J–3L were postulated as unstable intermediates yielding triol 4Avia benzaldehyde 5 and benzyl alcohol 3A intermediates. cis-Dihydroxylation of monosubstituted benzylic substrates containing bulky groups (1H, 1I) or 1,4-dialkyl-substituted benzene substrates (10A–10C) gave the corresponding cis-dihydrodiol metabolites (2H, 2I, 11A–11C) exclusively. The cis-diols 2A–2L, 11A–11C and triols 4A–4F, 4B′–4D′ were stereochemically assigned as single enantiomers of 1S,2R-configuration based on NMR and CD spectroscopy. The absolute configurations of the exocylic chiral centres in the triol bioproducts 4A–4F, 4B′–4D′ were established by stereochemical correlation and aromatisation/hydrogenation to yield the corresponding enantiopure phenolic benzylic alcohols having similar CD spectra.