Free radical intermediates formed during the oxidation of cyanide by horseradish peroxidase/H2O2 as detected with nitroso spin traps.

Aqueous solutions of cyanide react with hydrogen peroxide/horseradish peroxidase and form the cyanyl radical, which can be trapped by 2-methyl-2-nitrosopropane (t-nitrosobutane, tNB) at pH 9.8. At lower pH a variety of radical adducts are formed; at higher pH, the main product was the spin adduct of the formamide radical with tNB. The use of deuterated tNB and 15N-labeled potassium cyanide allowed the observation of the very small nitrogen coupling of this radical adduct. Experiments using 3,5-dibromo-4-nitrosobenzenesulfonic acid (DBNBS) as the spin trap yielded only the formamide radical adduct, which was identified by an independent synthesis starting from formamide. Both hydrogen splittings of its amino group could be resolved using deuterated DBNBS as the spin trap.