BACKGROUND: A spontaneous breathing trial (SBT) is commonly used to determine extubation readiness in patients receiving mechanical ventilation. However, the physiological impact of such a trial in preterm infants has not been well described. This study aimed to investigate the effects of a 3-min SBT on the cardiorespiratory stability of these infants. METHODS: A retrospective analysis of prospectively collected data was done for infants < 37 weeks gestational age who were extubated after a successful 3-min SBT. Heart rate, S pO 2 , breathing frequency, exhaled tidal volume, and Silverman Andersen Respiratory Severity Score (SA-RSS) to assess work of breathing, before and at the end of the SBT, were recorded and compared using nonparametric paired Mann-Whitney tests. A secondary analysis was done between extubation success (ie, 72 h without the need for re-intubation) and failure groups. Differences were considered statistically significant if P < .05. RESULTS: A total of 90 SBTs were performed in 70 premature infants; 65 had a successful SBT, and 5 failed the SBT. Of the 65 infants who had a successful SBT and were extubated, 6 failed extubation (9.2%). Subjects had a median (interquartile range [IQR]) gestational age of 30 (27-33) weeks at birth, a birthweight of 1,240 (860-1,790) g, and weight at extubation of 1,790 (1,440-2,500) g. Cardiorespiratory stability was noted by a significant decrease in median (IQR) exhaled tidal volume (6.4 [4.9-8.4] mL/kg vs 5.2 (3.8-6.6] mL/kg, P < .001), a significant increase in mean 6 SD breathing frequency (45.1 6 11.4 vs 52.6 6 14.4 breaths/min, P < .001), and a significant median (IQR) increase in work of breathing (SA-RSS of 1 [1-2] vs 2 [1-3], P < .001) at the end of the SBT. Respiratory instability was more remarkable in the success group. CONCLUSIONS: In preterm infants receiving prolonged mechanical ventilation, the performance of a 3-min SBT was associated with increased respiratory instability while still leading to a 10% extubation failure rate. Therefore, the routine use of SBTs to assess extubation readiness in this population is not recommended until there are clear standards and definitions, as well as good accuracy to identify failures.