Background <p>Effective management of preoperative anxiety is paramount in pediatric anesthesia, as untreated preoperative anxiety often leads to adverse clinical outcomes—most notably difficult parent-child separation and challenges during anesthesia induction. Therefore, preoperative sedation is essential for alleviating preoperative anxiety in preschool-aged children.</p> Methods <p>In this prospective randomized controlled clinical trial, children aged 2–6 years were assigned to one of two study groups: Group S (S-ketamine, 0.5&#xa0;mg/kg) and Group M (midazolam, 0.08&#xa0;mg/kg). The drugs were intravenously administrated in a 1:1 ratio in the anesthesia holding area. The primary outcome was the sedative effect, evaluated via the Modified Yale Preoperative Anxiety Scale (mYPAS). Second outcomes included the parent separation anxiety score, postoperative agitation, caregivers’ and anesthesia care providers’ satisfaction, and mask compliance.</p> Results <p>One hundred and four children were enrolled. Both prophylactic drugs effectively controlled preoperative stress. Compared with Group M, Group S had significantly lower mYPAS scores approximately 5&#xa0;min after intravenous premedication [median (interquartile range, IQR): 27.08 (22.92–33.33) versus 31.25 (27.08–33.33), <i>P</i> = 0.025]. A significant longer emergence time was found in Group S (<i>P</i> = 0.036). No significant differences were observed between the two groups in terms of parent separation anxiety score, mask compliance, or postoperative agitation.</p> Conclusion <p>Intravenous premedication with a single dose of 0.5&#xa0;mg/kg S-ketamine results in a deeper sedative state compared with 0.08&#xa0;mg/kg midazolam, albeit with a slight prolongation of emergence time.</p> Trial registration <p>Chinese Clinical Trial Registry (Registration number# ChiCTR2300069998); Date of Registration: 30/03/2023.</p>