The lipoxygenase pathway in tulip (Tulipa gesneriana):: detection of the ketol route

The in vitro metabolism of [1-C-14]linoleate, [1-C-14]linolenate and their 9(S)-hydroperoxides was studied in cell-free preparations from tulip (Tulipa gesneriana) bulbs, leaves and flowers. Linoleate and its 9-hydroperoxide were converted by bulb and leaf preparations into three ketols: (12Z)-9-hydroxy-10-oxo-12-octadecadienoic acid (alpha -ketol), (11E)-10-oxo-13-hydroxy-11-octadecadienoic acid (gamma -ketol) and a novel compound, (12Z)-10-oxo-11-hydroxy-12-octadecadienoic acid (10,11-ketol), in the approximate molar proportions of 10:3.1. The corresponding 15,16-dehydro alpha- and gamma -ketols were the main metabolites of [1-C-14]linolenate and its 9-hydroperoxide. Thus bulbs and leaves possessed 9-lipoxygenase and allene oxide synthase activities. Incubations with flower preparations gave alpha -ketol hydro(pero)xides as predominant metabolites. Bulb and leaf preparations possessed a novel enzyme activity, gamma -ketol reductase, which reduces gamma -ketol to 10-oxo-13-hydroxyoctadecanoic acid (dihydro-gamma -ketol) in the presence of NADH. Exogenous linolenate 13(S)-hydroperoxide was converted mostly into chiral (9S,13S)-12-oxo-10-phytodienoate (99.5% optical purity) by bulb preparations, while [1-C-14]linolenate was a precursor for ketols only. Thus tulip bulbs possess abundant allene oxide cyclase activity, the substrate for which is linolenate 13(S)-hydroperoxide, even though 13(S)-lipoxygenase products were not detectable in the bulbs. The majority of the cyclase activity was found in the microsomes (10(5) g pellet). Cyclase activity was not found in the other tissues examined, but only in the bulbs. The ketol route of the lipoxygenase pathway, mediated by 9-lipoxygenase and allene oxide synthase activities, has not been detected previously in the vegetative organs of any plant species.