Objective To determine the median effective dose (ED50) of remimazolam besylate in female patients with loss of consciousness. Methods A total of 32 women, American Society of Anesthesiologists (ASA) grade Ⅰ/Ⅱ, aged 35‒55 years old, who were scheduled to undergo hysteroscopy under total intravenous anesthesia were selected. During the induction of anesthesia, the patients were administered with remazolam besylate at one of the following five doses, 0.10, 0.13, 0.16, 0.20 mg/kg and 0.25 mg/kg in a sequential test. Then, ED50 and 95% effective dose (ED95) of remimazolam besylate, and corresponding 95% confidence intervals (CI) in female patients that caused unconsciousness were calculated by Probit regression analysis. Results The ED50 of remimazolam besylate in female patients that caused loss of consciousness was 0.175 (95%CI 0.153, 0.206) mg/kg, and ED95 was 0.255 (95%CI 0.213, 0.577) mg/kg. The breathing cycle was stable before and after administration of remimazolam besylate, and two patients had hiccups after administration. Conclusion According to the sequential test method, the ED50 of remimazolam besylate that caused loss of consciousness in female patients is 0.175 mg/kg.