DEVELOPMENT OF A CLASS-SPECIFIC COMPETITIVE ENZYME-LINKED-IMMUNOSORBENT-ASSAY FOR THE DETECTION OF PYRROLIZIDINE ALKALOIDS INVITRO

A two-step enzyme-linked immunosorbent assay (ELISA) was developed for the determination of the presence of retronecine in the parts per billion range. By modification of retronecine two haptens were synthesized. An I50 value of 11 +/- 3 ppb (Figure 2a) was obtained, with detection limits of 1.0-100 ppb. Indirect detection of monocrotaline (325 g/mol), retrorsine (351 g/mol), and senecionine (352 g/mol) was achieved by hydrolytic digestion of these naturally occurring pyrrolizidine alkaloids (PAs), demonstrating the utility of the assay as a class-specific detection method. PAs were detected in Senecio vulgaris and Crotalaria retusa plant sample extracts. No cross-reactivity was seen with either swainsonine or lupinine, both of which are structurally similar to retronecine in that they are also fused bicyclic alkaloids.