The burden of drug resistance emerges in the wake of chronic and repeated antibiotic use. This underpins the importance of discovering alternatives to current antibiotic regimens. In chronic rhinosinusitis (CRS), topical therapy such as nasal douches and steroid sprays is the mainstay of treatment. However, bacterial sinusitis such as those withStaphylococcus aureusbiofilm infection point to more recalcitrant CRS subtypes, focusing research efforts into topical antimicrobial therapies. In the sinuses, both local mucosal and systemic effects must be considered in designing any new topical medication. Nitric oxide (NO), an endogenous antimicrobial agent, is found at extremely low levels in CRS sinuses and high levels in healthy sinuses. As a novel treatment modality, we have designed a liposomal formulation of an NO donor (LFNO) using isosorbide mononitrate, as a topical sinus wash in a sheep model ofS. aureusbiofilm rhinosinusitis. Heart rate (HR), blood pressure, mean arterial pressure (MAP), and histologic and ciliary analyses were assessed in the safety component. Efficacy was assessed by quantifying biofilm biomass post-treatment. LFNO-treated sheep had lesser inflammation (P= 0.02), and comparable ciliary preservation (P= 0.86) than the control group. A transient increase in HR and decrease in MAP were observed in the LFNO group (P< 0.05), but this was not accompanied by observable side effects. LFNO sheep had significantly lower biofilm biomass vs controls (P= 0.044). Our findings demonstrate the localized and systemic safety of LFNO in an animal model despite using high NO concentrations, thus warranting further investigation for its possible therapeutic role in CRS.