Biosynthesis of the A/B/C/D-ring system of the rotenoid amorphigenin by Amorpha fruticosa seedlings

Abstract

With phenylalanine as the starting point, the biosynthesis of the characteristic rotenoid A/B/C/D-ring system of amorphigenin is studied using Amorpha fruticosa seedlings. The course of the biosynthesis can be divided into four phases represented by the bordered and interconnecting Schemes 1, 3, 6 and 7 which summarise the Chalcone–Flavanone Phase, the Flavanone–Isoflavone Phase, the Hydroxylation/Methoxylation Phase and the Rotenoid Phase. By using an INADEQUATE NMR experiment involving the administration of [1,2-¹³C]acetate, the type of folding forming ring-D is demonstrated by ¹³C–¹³C coupling and is interpreted as involving a polyketide containing a glutaconate segment which cyclises by a Claisen condensation. The resulting chalcone is cyclised, enzymically and stereospecifically, to 4′,7-dihydroxyflavanone.

The latter flavanone undergoes aryl migration, in a manner similar to that found in isoflavone biosynthesis, to give 7-hydroxy-4′methoxyisoflavone. Possible mechanisms for the flavanone–isoflavone rearrangement are discussed, including a proposal that the initiating step involves attack on ring-A and is similar to the first stage of the aromatic hydroxylation of tyrosine to dopa. Although possessing no 4′-hydroxy group in ring-A, the mechanism is also applicable to the recently discovered rotenoids of the Boerhaavia and Iris type, and it provides an explanation for the biogenesis of natural spirobenzocyclobutanes from dihydroeucominoids.

Six suitably substituted isoflavonoids labelled with ¹³C or ³H are synthesized and are used to show that the next hydroxylation (and probably methylation) involves C-3′ rather then C-2′ in 7-hydroxy-4′-methoxyisoflavone. Whilst the methylations involve S-adenosylmethionine, the hydro-xylating enzymes are probably very similar to the flavanone–isoflavone-rearranging enzyme. The closure of ring-B to form finally the rotenoid system probably involves conjugate addition of a methoxyl radical. Prenylation and oxidative modifications are characteristically late-stage processes.