Objective: The study objective was to evaluate the association of oral acetaminophen therapy versus immediate surgical ligation with neonatal outcomes in infants with persistent patent ductus arteriosus. Methods: We performed a retrospective cohort study of preterm infants born 28+6 weeks or less gestational age with persistent large patent ductus arteriosus being considered for surgical ligation after unsuccessful medical therapy. Infants in epoch 1 (July 2009 to June 2012) were immediately referred for ligation, and infants in epoch 2 (July 2012 to June 2015) were treated with oral acetaminophen and referred for ligation in the absence of improvement. The primary outcome was a composite of death or chronic lung disease. Results: A total of 92 infants with median (interquartile range) gestational age 25.2 weeks (24.4-26.3) had persistent large patent ductus arteriosus (43 in epoch 1, 49 in epoch 2). Infants in epoch 2 had decreased surgical ligation (26 [53%] vs 31 [72%]; adjusted odds ratio [aOR], 0.32; 95% confidence interval [CI], 0.12-0.89) but increased chronic lung disease (36 [73%] vs 25 [58%]; aOR, 3.34; 95% CI, 1.05-10.58) and increased death/chronic lung disease of borderline significance (39 [80%] vs 29 [67%]; aOR, 3.09; 95% CI, 0.99-9.63). Infants in epoch 2 took longer to wean off of positive pressure ventilation (28.5 vs 24 days after enrollment; aOR, 0.52; 95% CI, 0.31-0.85). Conclusions: Late oral acetaminophen therapy for infants with persistent patent ductus arteriosus is associated with reduced surgical ligation but increased chronic lung disease. In light of a lack of improvement in clinical outcomes, the individual contributory effects of acetaminophen, surgical ligation, and prolonged exposure to patent ductus arteriosus require further study to define the optimal approach.