Nitric oxide (NO)-releasing chitosan oligosaccharides were modified with ester functional groups to examine how the mucoadhesive nature of the scaffold impacts the ability of NO to degrade mucins from human bronchial epithelial cell cultures and clinical sputum samples collected from patients with cystic fibrosis (CF). Agarose gel electrophoresis experiments indicated that the mucoadhesive NO-releasing chitosan oligosaccharides degraded both the purified mucins and sputum, while control scaffolds (without NO release or mucoadhesive ligands) had no effect on mucin structure. Microscopic observations of sputum treated with the mucoadhesive NO-releasing chitosan oligosaccharide confirmed degradation of the mucin and DNA networks. Similarly, the viscosity and elasticity of sputum were reduced upon treatment with the mucoadhesive NO-releasing chitosan, demonstrating the potential utility of these NO-releasing scaffolds as mucolytic agents.