The aim of present study was to estimate the in vitro antioxidant activity of alcoholic extract of Cucumis callosus (Rottl.) Cogn (Cucurbitaceae) seeds which is commonly known as “Kachri” in Rajasthan (India) evaluated by using DPPH radical scavenging activity and hydrogen peroxide radical scavenging activity assay. The antioxidant activity was compared with ascorbic acid as standard. The IC50 values of Cucumis callosus and ascorbic acid were found 41.99 μg/ml and 24.27 μg/ml respectively for the DPPH radical scavenging activity while 95.27 μg/ml and 153.35 μg/ml respectively for hydrogen peroxide radical scavenging activity. Thus, alcoholic extract of Cucumis callosus seeds possess potent antioxidant activity in hydrogen peroxide model and may be useful for preparation of neutraceuticals as potent antioxidant to treat various human diseases. Key words: Cucumis callosus, Seeds, Antioxidant, Free radical scavenging, DPPH