Objective: Prosopis farcta has been used as a traditional herbal medicine for treating Diabetes mellitus. The aim of this study is to investigate the antidiabetic mechanisms of infusion (INF) extract of P. farcta and discovering the active extract for the first time. Materials and Methods: Six different extracts of P. farcta were prepared using five different solvents (ethanol, n-hexane, acetone, ethanol:water (1:1 v/v), and water). Cytotoxicity and cell proliferation assays were performed on mouse pancreatic β-cells (β-TC3) using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium method. The effects of P. farcta on glucose metabolism (in a hepatocellular carcinoma cell line [HepG2]) and glucose diffusion across a dialysis membrane (as a model of cellular glucose absorption) were evaluated. The protective effect of various P. farcta extracts on cytotoxicity, mitochondrial membrane potential (MMP), and streptozotocin (STZ)-induced apoptosis in β-TC3 cells was investigated. Results: Cytotoxicity study indicated that extracts were safe on β-TC3and HepG2 (≤0.5 mg/ml). INF protected β-TC3 cells from apoptosis induced by STZ and improved cell viability for 20% and significantly decrease depolarization of MMP (P < 0.005). The results showed that INF inhabited breaking/streaking the DNA. Proliferation study showed no significant increase in the number of cells either at single or multiple doses. In moderate hyperglycemia (11.1 mmol/l), a significant glucose-lowering effect was observed but glucose diffusion was not the probable mechanism of extracts antidiabetic effect. In conclusion, only INF, the traditionally used extract, has an antidiabetic potential by attenuating the death and apoptosis induced by STZ in β-TC3 cells and increase glucose consumption. Conclusion: The present study demonstrates that only INF extract have an antidiabetic potential by attenuating the death and apoptosis induced by STZ in β-TC3 cells and increase glucose consumption.