Oxidative stress plays an important role in the pathophysiology of bronchial asthma (BA), chronic obstructive pulmonary disease (COPD), and asthma-COPD overlap (ACO), but its relevance has not been fully elucidated. The aim of this study was to measure the levels of oxidative stress and investigate its clinical significance in patients with BA, COPD, or ACO. We recruited 214 patients between June 2020 and May 2023 (109 patients with BA, 63 with COPD, and 42 with ACO). To assess clinical conditions, we evaluated patient characteristics, results of respiratory function tests and blood tests, and administered several questionnaires. We evaluated oxidative stress using the test for derivatives-reactive oxygen metabolites (d-ROMs) in serum. The d-ROMs levels were significantly higher in patients with COPD or ACO than in patients with BA. There was no difference in serum d-ROMs levels between the COPD and ACO groups. In BA, d-ROMs levels were positively correlated with interleukin (IL)-6, IL-8, serum amyloid A (SAA), and C-reactive protein (CRP) levels; white blood cell (WBC) and neutrophil counts; and St. George's Respiratory Questionnaire (SGRQ) scores, and they were negatively correlated with forced expiratory volume in 1 s (%FEV) and asthma control test (ACT) score. In COPD, d-ROMs levels were positively correlated with IL-6, SAA, and CRP levels; WBC, neutrophil, and eosinophil counts; and COPD assessment test (CAT) and SGRQ scores, and they were negatively correlated with forced vital capacity (%FVC), %FEV, and %FEV/FVC scores. In ACO, d-ROMs levels were positively correlated with IL-6, SAA, tumor necrosis factor alpha (TNF-α), and CRP levels; and CAT and SGRQ scores, and they were negatively correlated with %FVC and %FEV scores. Serum d-ROMs levels may serve as a marker reflecting clinical conditions such as systemic inflammation, symptom severity, and airflow limitation in patients with BA, COPD, and ACO.