Cystic fibrosis patients demonstrate an increased susceptibility to bacterial lung infections. Airway infiltration by neutrophils will then lead to an increase in human leukocyte elastase (HLE) within the extracellular compartment, thereby producing deleterious effects. Here, we investigated the properties and tissue distribution of an unglycosylated, recombinant form of the HLE inhibitor alpha-1-proteinase inhibitor (alpha(1)-antitrypsin rhalpha1PI) when it is administered to the airway surface. We produced rhalpha1PI using a bacterial expression system and found the purified protein to be indistinguishable from blood-purified, glycosylated alpha1PI at inhibiting elastase in vitro. In contrast to intravenous administration, direct delivery of either alpha1PI or rhalpha1PI to the airway surface of CD-1 mice by nasal instillation produced similar highly detectable levels of protein in bronchoalveolar lavage at all time points, suggesting that glycosylation of alpha1PI does not play the same critical role in determining protein stability at the respiratory surface as it does in the vascular compartment. Interestingly, this unglycosylated rhalpha1PI was also highly protective against elastase-mediated injury 24 h after rhalpha1PI instillation and was consistently found to be significantly more protective than glycosylated blood-derived alpha1PI. Thus, these results provide evidence that aerosol delivery of rhalpha1PI could be an effective strategy for controlling HLE-dependent pathophysiology associated with cystic fibrosis lung disease.