Airway inflammation is a principal determinant of airway responsiveness and function in asthma and allergic diseases. Alveolar macrophages (AM) may contribute to inflammation in multiple ways, including release of reactive oxygen species such as superoxide (SO) anion. We hypothesized that SO production by AM increases after segmental bronchoprovocation (SBP) with relevant antigen and contributes to airway injury. Eight ragweed-sensitive subjects with allergic rhinitis were studied by bronchoalveolar lavage and ragweed SBP to determine the SO production and characteristics of cells recruited after antigen challenge. No significant changes in cell numbers or total protein concentration were observed immediately after antigen challenge. Purification (to greater than 94%) of AM on discontinuous gradients of Percoll revealed significantly increased spontaneous and opsonized-zymosan-driven SO production immediately after antigen challenge. Forty-eight hours later, total air-space cells, AM, eosinophils, and total protein concentration were significantly increased in relationship to antigen dose given. Furthermore, both unfractionated air-space cells and purified AM obtained 48 h after antigen challenge released increased amounts of SO anion in response to activator compared with either cells obtained immediately after SBP or those obtained 48 h after saline challenge. In addition, significant increases in high density AM were also seen 48 h after antigen challenge. These data suggest that AM activation occurs immediately after antigen challenge, and that the late airway response to antigen is characterized by the appearance of high density AM, which have potentiated SO release. The increased oxidative burden thereby produced may contribute to increased airway injury.