Synthesis of haptens and potential radioligands and development of antibodies to insect growth regulators diflubenzuron and BAY SIR 8514

A variety of synthetic approaches were undertaken, leading to potential haptens and radioligands for the benzoylphenylurea insect growth regulators diflubenzuron and BAY SIR 8514. One successful approach involved derivatization of the aniline nitrogen by ethyl 4-bromobutyrate followed by reaction with an appropriate isocyanate and cleavage of the ethyl ester to yield a free carboxypropyl "handle". Useful haptens were also synthesized by using a 3′-phenolic metabolite of diflubenzuron as well as acetate and amine functionalities in the 4′ position while the N-sulfenyl bond proved too unstable for use as an antigen. With the exception of the sulfenylated derivatives, the haptens lacked significant biological activity on three insect species. Following protein coupling by the active ester or water-soluble diimide method, antibodies were raised to two diflubenzuron haptens in each of seven rabbits immunized as demonstrated by radioimmunoassay using [¹⁴C]diflubenzuron, Ouchterlony gel diffusion, and immunoelectrophoresis.