Introduction:The seeds of Aegle marmelos (bael) are less explored for their therapeutic activity; cytotoxic activity of bael seeds was yet to be studied. The present study aims at finding the cytotoxic activity of the successive extracts and the flavonoid fraction from the methanolic extract of the seeds. Methods: The extracts were prepared using solvents (petroleum ether, chloroform, ethyl acetate, methanol, and water) in increasing order of their polarity. Anti-oxidant activity of the extracts was evaluated using DPPH radical scavenging assay, nitric oxide scavenging assay, and hydrogen peroxide scavenging assay. Flavonoid fractions were obtained by thin-layer chromatography of the flavonoid-rich fraction prepared from the total methanolic extract. Human colon cancer cell line (HT 29), human breast cancer cell line (MCF-7), human lung cancer cell line (A-549) were used to evaluate the cytotoxicity of the extracts and the flavonoid fractions. Results: Various phytochemicals like alkaloids, flavonoids, tannins and phenols, steroids, and saponins were detected in phytochemical investigation of the extract. Ethyl acetate fraction showed comparable DPPH radical scavenging activity and hydrogen peroxide scavenging activity with ascorbic acid (standard). Methanol extract showed good nitric oxide scavenging activity of all the extracts. Flavonoid fraction F-01 showed 50% cell growth inhibition at concentration <10 μg/ml on the human breast cancer cell line (MCF-7), which indicates its potent activity. Conclusion: F-01, one of the flavonoid fractions, showed comparable activity with the standard (Adriamycin) on MCF-7 cell line. Anti-oxidant study was conducted for the extracts, and the extracts showed good activity comparable to the standards used.