The current study was executed to isolate and evaluate gallic acid from Euphorbia hirta for in vitro radioprotective potentials against gamma irradiation caused radiotoxicity in human lymphocytes. The defatted E.hirta plant material was treated to methanol extraction using the soxhlet device. Bioflavonoids were isolated from the E.hirta methanol extract using column chromatography. In human cells exhibited to gamma radiation, separated flavonoid gallic acid was examined for in vitro radioprotective potentials using the micronucleus test, DNA fragmentation assay, superoxide free radical scavenging method, and apoptic assay. The frequency of micronuclei was considerably declined when cells were preprocessed with gallic acid (25g/mL) before being exhibited to 2Gy gamma radiation, as determined by the cytokinesis blocked micronucleus test. Similarly, pre-gamma radiation treatment of human cells with gallic acid led in markedly less DNA injury, as assessed by comet metrics like olive tail moment and percent tail DNA. Gallic acid (25g/mL) given to lymphocytes prior to gamma irradiation considerably decreased the percentage of apoptotic bodies. Gallic acid also considerably lowered the reactive oxygen species concentrations elicited by gamma radiation. Our findings showed that gallic acid protects lymphocytes isolated from human blood from gamma radiation-induced DNA destruction and anti-apoptotic activity, which could be because of inhibition of free radicals formed by gamma radiation as well as the decline of gamma radiation-induced oxidative stress.