Ageing is the process generated by senescent cells, free radicals, inflammation and other relevant factors. Ageing contributes to age-related diseases that affect the quality of life. People are interested in anti-ageing intervention and many scientists attempt to search for anti-ageing medicines. This review focused on describing in vivo anti-ageing activity of US-FDA-approved drugs and found that alogliptin, canagliflozin and metformin might produce anti-ageing activity via AMPK activation. Rapamycin and canagliflozin are capable to inhibit mTOR to promote lifespan. Atracurium, carnitine and statins act as DAF-16 activators, which potentially contribute to anti-ageing activity. Hydralazine, lisinopril, rosiglitazone and zidovudine may help stabilize genomic integrity to prolong life expectancy. Other indirect mechanisms, including insulin-lowering effect by acarbose and calcium channel blocking activity by verapamil may also promote longevity. Interestingly, some drugs (i.e., canagliflozin, metformin, rapamycin and acarbose) are likely to demonstrate a lifespan-promoting effect predominantly in male animals. These pre-clinical data might provide mechanistic and phenotypic perspectives to better understand the targets of anti-ageing interventions.