Background <p>To assess and compare the efficiency and outcomes between prone split-leg position and lithotomy position in posterior uterine myomectomy by transvaginal natural orifice transluminal endoscopic surgery (vNOTES).</p> Methods <p>33 patients with posterior uterine myomectomy by vNOTES in the prone split-leg position and 15 patients in the lithotomy position were retrospectively recruited. Important baseline characteristics and outcome parameters such as age, body mass index, volume of myoma, delivery mode, hospital length, intraoperative blood loss, hemoglobin values before and 72&#xa0;h after operation, VAS score, operation time and operation preparation time were compared between two patient groups.</p> Results <p>The operation time of the prone split-leg position group was significantly shorter than that of the lithotomy position group (<i>P</i> &lt; 0.05), but the operation preparation time of the prone split-leg position group was longer than that of the lithotomy position group (<i>P</i> &lt; 0.05). No significant difference was found in other indicators between the two patient groups.</p> Conclusions <p>Our study suggests the potential application of the prone split-leg position in posterior uterine myomectomy by vNOTES.</p>