Partially purified HCN Synthase (HCS) required exogenous electron accepters for activity. Phenazine methosulfate (PMS) provided the greatest activity, whereas oxygen allowed only a limited response. The P. aeruginosa secondary metabolite pyocyanin supported HCS-mediated cyanide production. HCN production by whole cells operated maximally at low oxygen levels, whereas moderate oxygen levels limited HCS activity. Respiration and cyanogenesis by whole cells were equally sensitive to azide; HCS was completely resistant.