© 2019, International Pediatric Research Foundation, Inc. Background: We evaluated the epidemiology of fluid balance (FB) over the first postnatal week and its impact on outcomes in a multi-center cohort of premature neonates from the AWAKEN study. Methods: Retrospective analysis of infants <36 weeks’ gestational age from the AWAKEN study (N = 1007). FB was defined by percentage of change from birth weight. Outcome: Mechanical ventilation (MV) at postnatal day 7. Results: One hundred and forty-nine (14.8%) were on MV at postnatal day 7. The median peak FB was 0% (IQR: −2.9, 2) and occurred on postnatal day 2 (IQR: 1,5). Multivariable models showed that the peak FB (aOR 1.14, 95% CI 1.10–1.19), lowest FB in first postnatal week (aOR 1.12, 95% CI 1.07–1.16), and FB on postnatal day 7 (aOR 1.10, 95% CI 1.06–1.13) were independently associated with MV on postnatal day 7. In a similar analysis, a negative FB at postnatal day 7 protected against the need for MV at postnatal day 7 (aOR 0.21, 95% CI 0.12–0.35). Conclusions: Positive peak FB during the first postnatal week and more positive FB on postnatal day 7 were independently associated with MV at postnatal day 7. Those with a negative FB at postnatal day 7 were less likely to require MV.