Objective To investigate the anatomy of anterior and posterior terminal perforators of the peroneal artery and its clinical applications. Methods Six lower limb specimens were obtained from 3 fresh cadavers. The anterior and posterior terminal perforators and the perforator of terminal peroneal artery were exposed under surgical microscope, and the distances from the beginning of each perforator branch to the lateral malleolus tip and the external diameter of each perforator were measured. With these anatomical knowledge and contrast-enhanced ultrasound (CEUS) guidance, the pedicle flaps with above-mentioned perforators were rationally selected and precisely designed for 18 patients with skin defects in the ankle and foot region between October 2016 and December 2018. Among the patients, there were 14 males and 4 females, aged 28-62 years, with an average age of 40 years. The area of wound ranged from 4 cm×3 cm to 13 cm×10 cm and the area of skin flap ranged from 5 cm×4 cm to 14 cm×10 cm. The anterior peroneal artery terminal perforator flap were applied in 13 cases and the posterior peroneal artery terminal perforator flap in 5 cases. The donor sites were closed directly in 7 cases and repaired with full thickness skin graft in 11 cases. Results The distance from the beginning of the anterior terminal perforator to the lateral malleolus tip was (5.1±0.5) cm, the external diameter of the anterior terminal perforator was (1.51±0.05) mm. The distance from the beginning of the posterior terminal perforator to the lateral malleolus tip was (4.9±0.9) cm, the external diameter was (1.78±0.17) mm; the distance from the beginning of the perforator of terminal peroneal artery to the lateral malleolus tip was (1.7±0.7) cm, the external diameter was (0.58±0.12) mm. Clinical application results: The edge of the flap was dark in 2 cases after operation and healed after surgical dressing, and 1 case of wound infection healed gradually after debridement. The other flaps survived and healed by first intention. Three patients underwent plastic surgery at 3 months after operation due to flap swelling. All patients were followed up 3-18 months. During the follow-up period, the flaps had good texture and appearance, and partial recovery of sensation. All cases were assessed by the American Orthopaedic Foot and Ankle Society (AOFAS) score at last follow-up. The results were excellent in 9 cases, good in 6 cases, fair in 2 cases, and poor in 1 case, with the excellent and good rate of 83.3%. ConclusionFurther classification of peroneal artery perforators in the lateral malleolus region can improve clinical understanding and be helpful to selection and application of perforator flaps in the lateral malleolus.