Abstract: Background and Objective: Macrotyloma uniflorum (MU) is claimed to be used in treatment of depression traditionally, in ayurveda, siddha and unani medicine. Also, the role of MU has potential source in curing depression is still unexplored. The present study was designed to evaluate the ethanolic extract of Macrotyloma uniflorum (EEMU) in experimental models of depression in rats. Materials and Methods: In the present study, phytochemical screening, including Thin-Layer Chromatography (TLC) and High-Performance Thin-Layer Chromatography (HPTLC) was performed to confirm the presence of isoflavone, daidzein, genisteins in standardized EEMU. The EEMU was screened at two doses (200 and 400 mg kg–1/p.o), based on acute toxicity study in rats. Each dose of EEMU was given to rats twice a day. Antidepressant activity was accessed by force swim test, tail suspension test and potentiation of nor-epinephrine toxicity. The duration of study was 1 week for all four groups. Antioxidants parameters like Thiobarbituric Acid Reactive Substances (TBARs), Reduced Glutathione (GSH) and nitrite/nitrate level were also accessed in brain homogenate to check the oxidative defence of EEMU extract. Results: The extract showed the presence of daidzein, genistein in EEMU. Further, study indicates that EEMU at doses (200 mg kg–1/p.o.) and (400 mg kg–1/p.o.) increase the mobility levels in rats as compared to control group. The treatment of rat with EEMU at a dose of (400 mg kg–1/p.o.) produced significant decrease in the levels of TBARs, nitrite/nitrate contents and increases the level of GSH that revealed the antioxidant nature of the extract. Conclusion: The antidepressant activity of EEMU due to its antioxidant and tyrosine kinase inhibiting nature.