Heavy metals such as mercury are some of the environmental pollutants and can induce toxicity by bioaccumulation and oxidative damage. This study aimed to investigate the effect of ethanolic extract of Medicago sativa L. (Alfalfa) on mercury damage in the kidney and liver of rats. Thirty Wistar rats were randomly divided into five groups, the control group, S group (2 mg/kg mercury chloride), and T1, T2, and T3 groups that, in addition to mercury, received doses of 250, 500, and 750 mg/kg of the alfalfa extract. On the last day, blood samples were taken, and the serum was separated to measure biochemical and oxidative stress parameters in the kidney and liver. A part of the kidney and liver was also used for histopathological evaluation. Total phenols and flavonoids were 40.45 ± 2.12 and 14.36 ± 0.45 mg/g, respectively, whereas IC50 was 245.18 ± 19.76 μg/ml. The body weight significantly decreased in the S group compared to other groups, while treatment with different doses of alfalfa extract increased the body weight. Mercury concentration in the kidney was higher than that in the liver. The serum levels of urea, creatinine, alanine aminotransferase (ALT), and alkaline phosphatase (ALP) significantly increased in the S group compared to the control group, while treatment with different doses of alfalfa extract increased their levels. Moreover, an increase in malondialdehyde (MDA) and a decrease in glutathione peroxidase (GPx), catalase (CAT), total antioxidant capacity (TAC), and superoxide dismutase (SOD) activity were observed in the S group. The level of these parameters significantly improved in the groups receiving the extract compared to the S group. Furthermore, the histopathological evaluation showed glomerular and tubular damage and hepatic necrosis in the S group and that these conditions improved in the T3 group. The findings of this study showed that the ethanolic extract of alfalfa in a dose-dependent manner has potentially unique protective effects against mercury poisoning in the kidney and liver.