Macrotermes barneyi Light, a fungus-growing termite widely distributed in southern China, cultivates fungus gardens composed mainly of fungal mycelia, soil, and partially digested plant fibers. These fungus gardens are nutrient-rich and represent a potential medicinal resource for developing novel health-promoting traditional Chinese medicine formulations. However, efficient extraction methods to maximize the antioxidant activity of M. barneyi fungus gardens remain unclear. In this study, fungus gardens were used as raw material, and the ABTS radical scavenging assay was employed to compare the antioxidant capacity of extracts obtained under different solid-liquid ratios (1∶4, 1∶5, 1∶10, 1∶15, 1∶20, 1∶25 (g/mL), ultrasonic time (5 min, 10 min, 20 min, 30 min, 40 min) and ultrasonic power (201.5 W, 305.5 W, 403 W, 500.5 W, 598 W). An orthogonal experimental design was used to optimize the ultrasonic extraction process. The results showed that the ABTS free radical scavenging capacity decreased with the decrease with a lower solid-liquid ratio. No significant difference was observed between the ratios of 1∶4 and 1∶5 g/mL. The scavenging capacity increased and then decreased with prolonged ultrasonic time, showing a bell-shaped trend, with the highest scavenging rate (38.75%) at 20 min. Similarly, scavenging capacity increased and then decreased with increasing ultrasonic power, peaking at 305.5 W with a scavenging rate of 20.28%. Orthogonal analysis indicated that the factors affecting ABTS radical scavenging capacity ranked in the order: solid-liquid ratio > ultrasonic power > ultrasonic time. The optimal extraction conditions were determined as a solid-liquid ratio of 1∶5 g/mL, ultrasonic time of 10 min, and ultrasonic power of 403 W, under which the ABTS radical scavenging rate reached 44.33%. This study shows that the effects of extraction parameters on the antioxidant activity of M. barneyi fungus gardens and identifies optimal conditions for ultrasonic extraction, providing a theoretical basis for the scientific utilization of termite fungus gardens and laying the groundwork for future studies on their active constituents and antioxidant mechanisms.