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Limonia acidissima L. (LA) is a fruit-bearing tree from the Rutaceae family that thrives in Tra Vinh province, Vietnam. It has been found that chemical compounds contained in LA leaves have antioxidant, antibacterial, anticancer, anti-inflammatory, and larvicidal properties. This study aimed to evaluate the antioxidant activity of leaf power ethanol-extracted using an ultrasound-assisted extraction technique. The input parameters of the process were optimized by the response surface methodology (RSM) including ethanol content (50 – 70 %), temperature (40 – 60 ºC), and time (30 – 50 min). The model response included total polyphenol content (TPC, Y1), and radical scavenging capacity by DPPH (Y2). The results show that the effect of the input parameters on the response of the extraction process was expressed by the quadratic equation with the high R2 values of Y1 (0.9827), and Y2 (0.9644). The optimal conditions for the extraction process were achieved at X1 = 59.7 %, X2 = 49.4 ºC, and X3 = 39.4 min. At this condition, the experimental value ??of TPC (85.79 ± 0.23 mg GAE/g dry weight, d.w) and DPPH (235.01 ± 0.79 µmol TEAC/g d.w) was not the significant difference compared to the predicted value. Furthermore, bioactive components from the leaf extract were quantified using modern GC-MS chromatography with the content of Stigmasterol (228.36 ± 4.95 ppm), Sitosterol (95.44 ± 3.3 ppm), Alpha-Pinene (60.40 ± 1.29 ppm). Antibacterial activity was tested against Escherichia coli, Salmonella enterica, Pseudomonas aeruginosa, and Staphylococcus aureus.