title: Direct evidence for membrane transport of host-plant-derived pyrrolizidine alkaloid N-oxides in two leaf beetle genera.
abstract: The chrysomelid leaf beetles Longitarsus jacobaeae, Oreina cacaliae, and O. speciosissima sequester pyrrolizidine alkaloids from their asteracean host plants and store them as nontoxic N-oxides. Previous analyses showed that Longitarsus is able to N-oxidize protoxic tertiary PAs, but did not resolve in which form N-oxides are taken up. For Oreina, beetles seem able to directly transmit the polar PA N-oxides from the gut into the hemolymph and prevent any reduction of them in the gut yielding protoxic free bases. Here, we confirm the predicted direct uptake of PAs as N-oxides by Oreina, and elucidate the situation for Longitarsus by applying double-labeled [14C]senecionine [18O]N-oxide as tracer. The beetles were fed with the tracer and subsequently senecionine N-oxide was recovered from the defensive secretions (Oreina) and beetle extracts (Longitarsus), purified by HPLC, and submitted to ESI-MS, GC-MS, and analysis of the specific radioactivity. The 18O-label is retained without any loss in the labeled senecionine N-oxide recovered from the two Oreina species. Analysis of the Longitarsus experiment was complicated by a contamination of the HPLC-purified senecionine N-oxide with a second compound, identified as a dihydrosenecionine N-oxide by high-resolution CID analysis. The dihydrosenecionine N-oxide, probably the 15,20-dihydro derivative, constitutes a major idiosyncratic senecionine metabolite present in the beetle. The recovered senecionine N-oxide retained 74% 18O-label. The remaining 25% is mostly due to loss of 18O by reduction and subsequent re-N-oxidation. The experiments confirm for both beetle genera a direct uptake of the polar nontoxic PA N-oxides, which requires specific membrane carriers. Accumulation of detrimental free base PA is prevented by glucosylation (Oreina) or N-oxidation (Longitarsus).
authors.affiliation: Biozentrum Grindel und Zoologisches Museum, Universität Hamburg, Martin-Luther-King-Platz 3, 20146 Hamburg, Germany. ingo.narberhaus@uni-hamburg.de
authors.forename: Ingo
authors.initials: I
authors.lastname: Narberhaus
authors.forename: Ulrich
authors.initials: U
authors.lastname: Papke
authors.forename: Claudine
authors.initials: C
authors.lastname: Theuring
authors.forename: Till
authors.initials: T
authors.lastname: Beuerle
authors.forename: Thomas
authors.initials: T
authors.lastname: Hartmann
authors.forename: Susanne
authors.initials: S
authors.lastname: Dobler
mesh_terms: D000818:Animals; D001692:Biological Transport; D001517:Coleoptera / Q000378:metabolism*; D003497:Cyclic N-Oxides / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D004435:Eating / Q000502:physiology; D008401:Gas Chromatography-Mass Spectrometry; D005960:Glucosides / Q000737:chemistry / Q000378:metabolism; D006458:Hemolymph / Q000378:metabolism*; D007553:Isotope Labeling; D008566:Membranes / Q000378:metabolism; D010084:Oxidation-Reduction; D011763:Pyrrolizidine Alkaloids / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*
chemical_list: D003497:Cyclic N-Oxides; D005960:Glucosides; D011763:Pyrrolizidine Alkaloids