A New Abscisic Acid Catabolic Pathway

We report the discovery of a new hydroxylated abscisic acid (ABA) metabolite, found in the course of a mass spectrometric study of ABA metabolism in Brassica napus siliques. This metabolite reveals a previously unknown catabolic pathway for ABA in which the 9ʹ-methyl group of ABA is oxidized. Analogs of (+)-ABA deuterated at the 8ʹ-carbon atom and at both the 8ʹ- and 9ʹ-carbon atoms were fed to green siliques, and extracts containing the deuterated oxidized metabolites were analyzed to determine the position of ABA hydroxylation. The results indicated that hydroxylation of ABA had occurred at the 9ʹ-methyl group, as well as at the 7ʹ- and 8ʹ-methyl groups. The chromatographic characteristics and mass spectral fragmentation patterns of the new ABA metabolite were compared with those of synthetic 9ʹ-hydroxy ABA (9ʹ-OH ABA), in both open and cyclized forms. The new compound isolated from plant extracts was identified as the cyclized form of 9ʹ-OH ABA, which we have named neophaseic acid (neoPA). The proton nuclear magnetic resonance spectrum of pure neoPA isolated from immature seeds of B. napus was identical to that of the authentic synthetic compound. ABA and neoPA levels were high in young seeds and lower in older seeds. The open form (2Z,4E)-5-[(1R,6S)-1-Hydroxy-6-hydroxymethyl-2,6-dimethyl-4-oxo-cyclohex-2-enyl]3-methyl-penta-2,4-dienoic acid, but not neoPA, exhibited ABA-like bioactivity in inhibiting Arabidopsis seed germination and in inducing gene expression in B. napus microspore-derived embryos. NeoPA was also detected in fruits of orange (Citrus sinensis) and tomato (Lycopersicon esculentum), in Arabidopsis, and in chickpea (Cicer arietinum), as well as in drought-stressed barley (Hordeum vulgare) and B. napus seedlings.