Unilateral sinus disease (USD) can sometimes be difficult to accurately diagnose before surgery. The application of nasal nitric oxide (nNO) for USD diagnosis and its surgical outcome in USD has not been reported in the literature. We prospectively enrolled sixty-six USD patients who underwent endoscopic sinus surgery for fungal rhinosinusitis (n = 19), chronic rhinosinusitis (CRS) without nasal polyps (n = 13), CRS with nasal polyps (n = 12) and sinonasal mass lesions (n = 22). nNO levels were measured preoperatively and at three and six months postoperatively. Correlations between nNO levels and potential clinical parameters, type of disease, disease severity, and disease-related quality of life (QOL) were assessed. Unlike bilateral CRS, in USD, nNO levels did not correlate with disease severity or postoperative QOL improvements. Except for fungus group, there were no differences in nNO levels between lesion and non-lesion sides in all the other groups. nNO levels on both sides were significantly elevated six months postoperatively in all groups. Fungal rhinosinusitis patients had the lowest preoperative nNO levels, and a cutoff of 239.3 ppb had the best sensitivity (79.0%) and specificity (87.2%) for preoperative diagnosis. While preoperative nNO levels cannot serve as an alternative marker for disease severity of USD, they were lower in fungal rhinosinusitis patients than in other USD patients and may be useful for more accurate diagnosis prior to surgery.