An extraordinary accumulation of (−)-pinoresinol in cell-free extracts of Forsythia intermedia: evidence for enantiospecific reduction of (+)-pinoresinol

Abstract

Stereoselective and enantiospecific transformation mechanisms in lignan biogenesis are only now yielding to scientific inquiry: it has been shown that soluble cell-free preparations from Forsythia intermedia catalyse the formation of the enantiomerically pure lignan, (−)-secoisolariciresinol, when incubated with coniferyl alcohol in the presence of NAD(P)H and H 2O 2. Surprisingly, (−)-pinoresinol also accumulates in this soluble cell-free assay mixture in > 96 % enantiomeric excess, even though it is not the naturally occurring antipode present in Forsythia sp. But these soluble cell-free preparations do not engender stereoselective coupling; instead, racemic pinoresinols are first formed, catalysed by an H 2O 2-dependent peroxidase reaction. An enantiospecific NAD(P)H reductase then converts (+)-pinoresinol, and not the (−)-antipode, into (−)-secoisolariciresinol. Stereoselective syntheis of (+)-pinoresinol from E-coniferyl alcohol is, however, catalysed by an insoluble enzyme preparation in F. suspensa, obtained following removal of readily soluble and ionically bound enzymes; no exogenously supplied cofactors were required other than oxygen, although the reaction was stimulated by NAD-malate addition. Thus, the overall biochemical pathway to enantiomerically pure (−)-secoisolariciresinol has been delineated.