Astragalus L. (Fabaceae) is an important genus with numerous species having various traditional medicinal uses making them of interest for scientific investigations to ascertain their therapeutic benefits. In the present study, the quantitative polyphenolic profiles of methanolic extracts from different parts (leaves, flowers, and roots) of two endemic Astragalus species growing in Turkey, i.e. A. campylosema Boiss. and A. hirsutus Vahl were determined, along with their antioxidant and enzyme inhibitory properties. A. campylosema and A. hirsutus extracts showed varying total phenolic (25.80–40.60 and18.59–29.46 mg GAE/g, respectively) and total flavonoid (11.21–105.91 and 16.06–131.91 mg RE/g, respectively) contents. HPLC-MS/MS revealed rutin to be the predominant phenolic compound in all the extracts of A. campylosema and leaf extract of A. hirsutus (133.53–752.42 μg g−1), while hyperoside was the major one in the flower and root extracts of A. hirsutus (2014.07 and 123.13 μg g−1, respectively). In DPPH and ABTS assays, radical scavenging capacity was demonstrated by all extracts of A. campylosema (47.13–48.10 and 87.03–115.36 mg TE/g, respectively) and A. hirsutus (17.82–38.67 and 47.84–57.29 mg TE/g, respectively). Reducing activity was also displayed by the extracts in CUPRAC and FRAP assays (A. campylosema: 83.06–135.20 and 59.15–90.19 mg TE/g, respectively; A. hirsutus: 53.02–83.42 and 31.25–43.25 mg TE/g, respectively). All extracts were also found to act as metal chelators (12.32–21.45 mg EDTAE/g) and exhibited total antioxidant capacity ranging from 1.16 to 1.60 mmol TE/g, in phosphomolybdenum assay. Acetyl- and butyryl-cholinesterase inhibitory effects were observed by all the extracts of the two species (1.56–4.99 mg GALAE/g). Anti-hyperpigmentation potential by inhibiting tyrosinase (54.55–67.35 mg KAE/g) was reported as well. Carbohydrate hydrolyzing enzymes, amylase and glucosidase were also inhibited (0.22–1.03 mmol ACAE/g). Overall, A. campylosema extracts showed relatively better antioxidant and enzyme inhibitory potentials compared to A. hirsutus extracts. Strikingly, A. hirsutus extracts was found to have higher AGE inhibition activity than A. campylosema. Although the cytotoxic effect of three different organs obtained from A. campylosema and A. hirsutus increased depending on the dose (from 10 to 200 μg/mL), it was found that both plant extracts did not show a genotoxic effect at the highest concentration of 200 μg/mL. Indeed, data amassed from this current scientific work showed the two selected Astragalus species to be rich in bioactive polyphenols that could be responsible for the various pharmacological activities and hence demands to be further explored for their possible applications as natural health promoting agents.