Several species of Longitarsus take up, metabolize and store pyrrolizidine alkaloids (PAs) from their host plants. In feeding experiments using radioactively labeled PAs of different types we examined the time course of the sequestration process in L. jacobaeae and L. aeruginosus. We found that adapted species efficiently store PAs for at least two weeks without major losses. During that time, there is virtually no change in the ratio of tertiary alkaloids to stored non-toxic N-oxides, regardless of chemical form fed to the beetles. This implies a transient N-oxidation process where the alkaloids are only temporarily accessible to the enzyme. A dissection experiment with L. aeruginosus six days after uptake of labeled PAs showed that the tertiary alkaloids are not found in the hemolymph but are stored in the elytra and other body compartments. This conforms with earlier experiments that localized the enzyme’s site of action in the hemolymph. Furthermore we show that different total alkaloid doses in the diet of L. jacobaeae and the potentially less adapted L. succineus do not affect the ratio of recovered N-oxides to tertiary molecules. Thus, the efficiency of the N-oxidizing enzyme is not dependent on the concentration of alkaloids offered.