Our early work indicated that methanolic extracts from the flowers, leaves, bark, and isolated compounds ofAcacia salignaexhibited significant antioxidant activities in vitro. The overproduction of reactive oxygen species (ROS) in the mitochondria (mt-ROS) interfered with glucose uptake, metabolism, and its AMPK-dependent pathway, contributing to hyperglycemia and diabetes. This study aimed to screen the ability of these extracts and isolated compounds to attenuate the production of ROS and maintain mitochondrial function via the restoration of mitochondrial membrane potential (MMP) in 3T3-L1 adipocytes. Downstream effects were investigated via an immunoblot analysis of the AMPK signalling pathway and glucose uptake assays. All methanolic extracts effectively reduced cellular ROS and mt-ROS levels, restored the MMP, activated AMPK-α, and enhanced cellular glucose uptake. At 10 µM, (−)-epicatechin-6(from methanolic leaf and bark extracts) markedly reduced ROS and mt-ROS levels by almost 30% and 50%, respectively, with an MMP potential ratio 2.2-fold higher compared to the vehicle control. (−)-Epicatechin6increased the phosphorylation of AMPK-α by 43%, with an 88% higher glucose uptake than the control. Other isolated compounds include naringenin1, naringenin-7-O-α-L-arabinopyranoside2, isosalipurposide3,D-(+)-pinitol5a, and (−)-pinitol5b, which also performed relatively well across all assays. AustralianA. salignaactive extracts and compounds can reduce ROS oxidative stress, improve mitochondrial function, and enhance glucose uptake through AMPK-α activation in adipocytes, supporting its potential antidiabetic application.