SYNTHESIS OF NITROXIDES FOR USE AS PROCATIONIC LABELS AND THEIR INCORPORATION INTO NAFION FILMS

A nitroxide label was covalently attached to amphetamine and two antid epressants, nortriptyline and desipramine, by forming an amide linkage between the amino group on each of these compounds and 3-carboxyl-2,2 ,5,5-tetramethyl-1-pyrrolidinyloxy. This nitroxide label was also atta ched to be biologically important compound biotin, but a spacer group, N,N'-dimethyl-1,6-hexanediamine, was needed to make two amide linkage s between the carboxylic acids on the nitroxide and biotin. These nitr oxide-labeled substances undergo a one-electron reversible oxidation b etween 0.5 and 0.6 V (vs. Ag/AgCl) at a naked glassy carbon (GC) elecr ode and between 0.3 and 0.4 V at a GC electrode coated with a polyanio nic film of Nafion. When a 0.8-V potential is applied to a GC/Nafion e lectrode in the presence of an aqueous buffer solution of one of these nitroxides, the nitroxide preconcentrates in the film in its oxidized form, the oxoammonium cation. Subsequent scanning of the potential in the negative direction using square wave voltammetry produces a reduc tion wave with a relatively large peak current making it possible to d etect almost nanomolar quanitities of the procationic nitroxide. This preconcentration of procationic nitroxides makes this redox label usef ul in a recently developed analytical technique that combines immunoas say with Nafion-modified electrodes.