Formation of a new class of oxylipins from N-acyl(ethanol)amines by the lipoxygenase pathway.

N-Acylethanolamines (NAEs) constitute a new class of plant lipids and are thought to play a role in plant defense strategies against pathogens. In plant defense systems, oxylipins generated by the lipoxygenase pathway are important actors. To date, it is not known whether plants also use endogeneous oxylipins derived from NAEs in their defense reactions. We tested whether members of the NAE class can be converted by enzymes constituting this pathway, such as (soybean) lipoxygenase-1, (alfalfa) hydroperoxide lyase and (flax seed) allene oxide synthase. We found that both alpha-N-linolenoylethanolamine and gamma-N-linolenoylethanolamine (18:3), as well as alpha-N-linolenoylamine and gamma-N-linolenoylamine were converted into their (13S)-hydroperoxide derivatives by lipoxygenase. Interestingly, only the hydroperoxides of alpha-N-linolenoyl(ethanol)amines and their linoleic acid analogs (18:2) were suitable substrates for hydroperoxide lyase. Hexanal and (3Z)-hexenal were identified as volatile products of the 18:2 and 18:3 fatty acid (ethanol)amides, respectively. 12-Oxo-N-(9Z)-dodecenoyl(ethanol)amine was the nonvolatile hydrolysis product. Kinetic studies with lipoxygenase and hydroperoxide lyase revealed that the fatty acid ethanolamides were converted as readily or even better than the corresponding free fatty acids. Allene oxide synthase utilized all substrates, but was most active on (13S)-hydroperoxy-alpha-N-linolenoylethanolamine and the (13S)-hydroperoxide of linoleic acid and its ethanolamine derivative. alpha-Ketols and gamma-ketols were characterized as products. In addition, cyclized products, i.e. 12-oxo-N-phytodienoylamines, derived from (13S)-hydroperoxy-alpha-N-linolenoylamines were found. The results presented here show that, in principle, hydroperoxide NAEs can be formed in plants and subsequently converted into novel phytooxylipins.