Objective: To determine if high frequency oscillatory ventilation (HFOV) decreases surfactant production in premature infants with respiratory distress syndrome (RDS). Study design: We randomized 19 infants <28 weeks of gestation to either HFOV (n = 8) or conventional ventilation (CV, n = 11) at 24 hours of life. After a 24-hour continuous infusion of uniformly labeled carbon 13 glucose (U∼13C6) glucose, we measured 13C enrichment in surfactant phosphatidylcholine (PC) in tracheal aspirate samples using gas chromatography/mass spectrometry. We calculated the fractional synthetic rate (FSR) of surfactant PC from labeled glucose and its half-life of clearance (T1/2). Results: FSR did not differ between groups (4.7% ± 2.7%/day CV vs 4.2% ± 3.1%/day HFOV, P = .7). T1/2 was 79 ± 18 hours in the CV group and 76 ± 23 hours in the HFOV group (P = .7). Neither degree of ventilatory support nor supplemental oxygen exposure correlated with surfactant metabolic indices. Three of 4 infants who died from RDS within the first month of life had a shorter T1/2 than 14 of 15 infants who survived. Conclusion: Surfactant metabolism is similar in preterm infants ventilated with HFOV and CV. Shortened surfactant half-life may characterize a subset of preterm infants with lethal RDS.