Electrochemical nitric oxide (NO) reduction by heme groups incorporated in films of didodecyldimethylammonium bromide (DDAB) on pyrolitic graphite was investigated. It is shown that DDAB most likely induces the release of the heme group from myoglobin and therefore myoglobin-DDAB and heme-DDAB films give the same voltammetric responses. This is confirmed by UV/vis spectroscopy showing a clear shift in the Soret band of myoglobin in a DDAB solution. The electrochemical NO reduction on a heme-DDAB film at different pH values reveals the presence of pH-dependent and pH-independent NO reduction pathways. The selectivity of these pathways is probed by combining the rotating ring-disk electrode technique with online electrochemical mass spectroscopy showing that the product of the pH-independent pathway is N2O and the product of the pH-dependent pathway is NH2OH. The preference for one or the other pathway seems to depend on whether a proton or a NO molecule is transferred to a Fe-II-NO- reaction intermediate and is influenced by pH, NO concentration, and potential.