Background: Hyperthyroidism can be a serious health problem, if not treated properly. This investigation primarily aimed to evaluate the thyroid regulatory and hepatoprotective activities of ethyl acetate fraction (EPBL) of Piper betle leaf extract in L-thyroxine (L-T4)-induced hyperthyroid rats. Materials and Methods: Effects of EPBL extract at a prestandardized dose of 50 mg/kg (p. o.) were studied in L-T4 (500 μg/kg/d, i. p.)-administered rats to examine the alterations in the levels of serum triiodothyronine (T3), thyroxine (T4), thyrotropin, alanine transaminase, and aspartate aminotransferase; in the activities of hepatic 5'-monodeiodinase (5'DI), glucose-6-phosphatase (G-6-Pase), and Na+-K+-ATPase; in the level of tissue malondialdehyde (MDA) and lipid hydroperoxides (LOOHs); and in the activities of antioxidants. Results: Administration of the EPBL extract reversed the T4-induced increase in serum thyroid hormones, the liver marker enzymes, MDA, and LOOH but enhanced the activities of antioxidative enzymes and reduced the glutathione content. Light microscopic findings of liver histology revealed distorted hepatic tissue architecture in hyperthyroid animals that were improved by the EPBL administration. High-resolution liquid chromatography–mass spectroscopy analysis revealed four main flavonoid glycosides such as quercetin, rutin, kaempferol, and luteolin. Conclusion: For the first time, our findings revealed the antithyroidic property of EPBL in T4-induced hyperthyroidism, without any hepatotoxicity. The antithyroidic and antioxidative properties of EPBL in hyperthyroid animals could be due to the presence of flavonoid glycosides in the extract which might have inhibited the thyroid hormone secretion and conversion of T4 to T3 through an inhibition of 5'DI.