The endangered Persian onager (Equus hemionus onager) has experienced significant population declines over the past century due to poaching, habitat destruction, and resource competition. Remaining animals in zoos and the wild are regionally isolated. Artificial insemination (AI) may be particularly useful as a means of aiding in global genetic management of these isolated populations. The first successful AI in onagers was performed in 2009 utilizing urinary hormone analyses and regular transrectal ultrasound examinations that required specialized handling devices. A method for estrous synchronization in this species would alleviate the need for daily handling and provide a more feasible approach to AI. This study tested long-acting controlled-release preparations of estradiol and progesterone, followed by a single injection of prostaglandin 10 days later, in six adult female Persian onagers to determine whether ovulation would occur within a narrow window of time. Serial transrectal ultrasound exams were performed to determine the day of ovulation following hormone treatment. Means and standard deviations were determined for the lengths of follicular and luteal phases, follicle sizes, and time to ovulation, and compared to historical data in this species. All six onagers ovulated between Days 18 and 22, with three females ovulating on Day 19, as determined by the presence of a corpus luteum. This is an apparently safe and effective method for the synchronization of estrous cycles in the Persian onager, and may be used to develop a timed AI protocol for use at institutions that do not have specialized handling facilities to enable regular transrectal ultrasound.