Pulmonary inflammation in asthma is orchestrated by the activity of NF-κB. NO and NO synthase (NOS) activity are important modulators of inflammation. The availability of the NOS substrate, l-arginine, is one of the mechanisms that controls the activity of NOS. Arginase also uses l-arginine as its substrate, and arginase-1 expression is highly induced in a murine model of asthma. Because we have previously described that arginase affects NOx content and interferes with the activation of NF-κB in lung epithelial cells, the goal of this study was to investigate the impact of arginase inhibition on the bioavailability of NO and the implications for NF-κB activation and inflammation in a mouse model of allergic airway disease. Administration of the arginase inhibitor BEC (S-(2-boronoethyl)-l-cysteine) decreased arginase activity and caused alterations in NO homeostasis, which were reflected by increases in S-nitrosylated and nitrated proteins in the lungs from inflamed mice. In contrast to our expectations, BEC enhanced perivascular and peribronchiolar lung inflammation, mucus metaplasia, NF-κB DNA binding, and mRNA expression of the NF-κB-driven chemokine genes CCL20 and KC, and lead to further increases in airways hyperresponsiveness. These results suggest that inhibition of arginase activity enhanced a variety of parameters relevant to allergic airways disease, possibly by altering NO homeostasis.