Aims andObjectives: To analyze stress levels, sleep quality, sleepiness and chronotypes on emergency room (ER) professionals before and during the COVID-19. Background: ER healthcare professionals are exposed to high stress levels, and they often present with poor sleep quality. Design: Observational study conducted into two different phases (before the COVID-19 and during the first wave of the COVID-19). Method: Physicians, nurses and nursing assistants working in the ER were included. Stress, sleep quality, daytime sleepiness and chronotypes were assessed by the Stress Factors and Manifestations Scale (SFMS), the Pittsburgh Sleep Quality Index (PSQI), the Epworth Sleepiness Scale (ESS), and the Horne and Österberg Morningness-Eveningness questionnaire, respectively. The first phase of the study was performed between December 2019 and February 2020, and the second between April and June 2020. The STROBE checklist was used to report the present study.Results: In total, 189 ER professionals in the pre-COVID-19 phase and 171 (of the initial 189) during COVID-19 were included. The proportion of workers with a morning circadian rhythm increased during the COVID-19, and stress levels were significantly higher during COVID-19 compared to the previous phase (38.34 ± 10.74 vs. 49.97 ± 15.81). ER professionals with poor sleep quality presented higher stress in