Overdose of acetaminophen causes hepatotoxicity due to NAPQI formation. The green synthesis of gold nanoparticles represents as a novel drug carrier in the field of drug delivery system. This study was designed to investigate the protective effect of green synthesized herbal gold nanoparticles (AuNPs) using the aqueous bark extract of Terminalia arjuna against acetaminophen-induced hepatotoxicity in an experimental rat model. In this study, group 1 served as normal control; group 2 received acetaminophen intraperitoneally at a concentration of 500 mg/kg of body weight for 14 days; and groups 3, 4, 5, and 6 were co-administered with acetaminophen (500 mg/kg/day) and AuNPs (55, 175, 550, 2000 μg/kg/day) intraperitoneally for 14 days. After 14 days, all animals were sacrificed for biochemical and histopathological studies. Among different experimental doses of AuNPs (55, 175, 550, 2000 μg/kg/day), dose 175 μg/kg/day showed more potent activity towards wellness parameters, biochemical indices, and histopathological studies. There was a significant (p < 0.05) increase in SGOT, SGPT, ALP, bilirubin, and MDA levels, but a significant decrease in SOD, CAT, and GSH activities in the hepatotoxic group in comparison with the control group, but co-administration with AuNPs (175 μg/kg/day) restored the activities of these biochemical markers. Hence, this study confirmed that AuNPs at a dose 175 μg/kg/day has better hepatoprotective efficacy. © 2020, Springer Science+Business Media, LLC, part of Springer Nature.