Timosaponin AⅢ(TAⅢ), derived from the Chinese medicinal herb Anemarrhena asphodeloides Bunge, has been reported to have a range of pharmacological effects including improvement of learning and memory deficits, anti-tumor, hypoglycemic effect and anti-hypertension. This study explored the therapeutic effects and preliminary mechanisms of TAⅢ in improving insulin resistance in ob/ob mice. We found that treatment with 10 mg kg-1·d-1 of TAⅢ reduced the expression of SREBPs and alleviated ectopic lipid deposition by decreasing DAG accumulation in liver. The decrease of DAG further inhibited the membrane translocation of PKC-ε, releasing its inhibition of phosphorylation at Ser307 of IRS1, and ultimately enhancing the AKT signaling response to insulin stimulation. In addition, TAⅢ promoted the browning of iWAT by activating the PGC1α-UCP1 axis on ob/ob mice, thereby enhancing fatty acid oxidation and increasing energy consumption, thus reducing its interference with insulin signaling. TAⅢ worked by enhancing the function of adipose tissue and inhibited lipid synthesis. These actions collectively ameliorated metabolic disturbances associated with insulin resistance. Therefore, we preliminarily concluded that TAⅢ improved metabolic disturbances related to insulin resistance. However, further research is needed, additional studies are necessary to validate these potential mechanisms.