Biosynthesis of defensive allomones in leaf beetle larvae: Stereochemistry of salicylalcohol oxidation in Phratora vitellinae and comparison of enzyme substrate and stereospecificity with alcohol oxidases from several iridoid producing leaf beetles

(7S)-[H-2(5)]-Salicylalcohol (3) and (7R)-[H-2(1)]-salicylalcohol (5) have been synthesized in order to examine the stereospecificity of salicylalcohol oxidase from the defensive secretion of the salicylaldehyde-producing leaf beetle Phratora vitellinae. Oxidation was found to proceed by selective removal of the C(7)-H-R hydrogen atom (Re-specificity) to yield salicylaldehyde. (7S)-[H-2(6)]-Benzylalcohol (9) was also oxidized Re-specifically to benzaldehyde, but in much lower yield, indicating the importance of the orthohydroxy group of salicylalcohol in substrate enzyme binding. The stereospecificities of terpenoid oxidases from six species of iridoid-producing leaf beetle were examined using (1R,8R)-[H-2(2)]-8-hydroxygeraniol (10), and were all found to oxidize the substrate Re-specifically. Cross-activity of oxidation was found in a number of species, with P. vitellinae able to oxidize terpenoid (10) and two of the iridoid-producing species able to oxidize salicylalcohol analogue (3), again with Re-specificity. However, when the two substrate analogs were presented together, in equal concentrations, preferential oxidation of the natural analog was observed in each case. The kinetics of oxidation for a number of terpenoid and aromatic alcohols by the defensive secretion of the iridoid-producing leaf beetle Phaedon armoraciae have been studied, revealing a large difference between the rate of(primary, allylic) terpenoid alcohol oxidation and the rate of salicylalcohol oxidation, thus accounting for the observed selectivity.