Biosynthesis of canescin, a metabolite of Aspergillus malignus: incorporation of methionine, acetate, succinate, and isocoumarin precursors, labelled with deuterium and carbon-13

Abstract

The biosynthesis of canescin (1), a metabolite of Aspergillus malignus, has been studied using [Me¹³C²H₃] methionine, sodium [1,2-¹³C₂]- and [1-¹³C, 2-²H₃]acetate, and sodium [2,3-¹³C₂]- and [2-¹³C, 2-²H₂]-succinate as simple precursors, and deuterium labelled isocoumarins as potential advanced precursors. Analysis of the labelled products by both n.m.r. and mass spectrometry suggests that 6,8-dihydroxy-3,7-dimethylisocoumarin (13) is the first enzyme-free intermediate produced by the polyketide synthase, and the 7-formyl-6,8-dihydroxy-3-methylisocoumarin (16) is a later intermediate on the pathway. Incorporation into canescin of ¹³C together with one deuterium atom from [¹³C²H₃]methionine proves that the formyl group of (16) is derived from this source and that it is not oxidised to the level of a carboxylic acid in subsequent steps. The ¹³C labels in the succinate units are incorporated into the γ-lactone carbons (C-11 to C-13) of canescin, but with complete loss of the neighbouring deuterium label.