Caragana ambigua stocks has been used in different traditional medicinal systems to treat diabetes and various stomach disorders. It has also been utilized for the nourishment and the regulation of blood flow. However, the biological profile and phytochemistry of this plant still need to be explored. The objective of current study is to probe therapeutic potential of Caragana ambigua. Methanolic extract of Caragana ambigua and its five fractions namely, n-hexane, chloroform, ethyl acetate, n-butanol and aqueous were evaluated for in vitro antioxidant (total phenolic and flavonoid content, DPPH assay, DNA protection assay), antidiabetic (α-amylase inhibition assay), antimicrobial (disc diffusion assay) and cytotoxicity studies (XTT assay and RBC’s cellular membrane protection assay). Amongst the investigated fractions and methanolic extract, the highest phenolic content (80 mg/g gallic acid equivalent) was observed in n-hexane and chloroform fraction whilst the maximum flavonoid content (18 mg/g quercetin equivalent) and the most significant (p<0.05) α-amylase enzyme inhibition was also displayed by n-hexane fraction (IC₅₀ = 252.2±1.08 μg/mL). Whereas the significant (p<0.05) DPPH inhibition was found in butanol fraction (IC₅₀ = 76.44±1.50 μg/mL). Moreover, butanol fraction at 200 μg/mL was also observed to protect the DNA from nicking. Methanolic extract and its analyzed fractions also exhibited cytotoxicity potential against human breast cancer and T-lymphoblastic leukemia cell lines. However, significant (p<0.05) dose dependent inhibitory effects on cell growth were observed against T-lymphoblastic leukemia cell lines where the maximum cytotoxicity (7.533±0.430%) was exhibited by n-hexane fraction. No substantial findings were obtained in case of antimicrobial activity against the tested strains. Lastly, hemolytic potential was found to be in the safe range. Thus, it can be concluded that Caragana ambigua can be exploited as an alternative approach to treat diabetes, cancer and other diseases by reducing oxidative stress.