Many components in Crotalaria assamica benth composition make this plant a potential antioxidant and antibacterial herbal material, which can find many applications in medicine and biochemistry. In this study, Crotalaria assamica benth seed powder was extracted with aqueous alcohol solutions by an ultrasound-assisted leaching method. The ultrasound wave was 37 kHz, and the extraction was carried out at 30 °C for 10 mins. An alcohol content of 54.78 vol% and a mass ratio of solid to liquid (RSL) of 1/55 are suitable conditions according to the reaction surface method (RSM) developed by the Box–Beknen approach to design matrix experiments. With the above conditions, the actual result was that the TPC was 12.02 ± 0.24 mg GAE/g DW and the TFC was 7.68 ± 0.34 mg RE/g DW). Crotalaria assamica confers antioxidant activity of the extract with an IC50 value of 644 ??g/mL, and antibacterial activity against P. aeruginosa strain, with an average antibacterial ring diameter of 13.5 mm.