Traditionally known as Pashanabheda in India, Aervalanata (L) belonging to the family Amaranthaceae, is widely available in Western ghats of India and used as Antiurolithiatic, Antimicrobial, Antiinflammatory, Hepatoprotective drug by Indian traditional system of medicines. Materials and Method: In the present study this herb is subjected to extraction with Pet. Ether, Chloroform, Ethyl acetate and Methanol followed by fractionation with different solvents of varying polarities such as n-hexane: Acetic acid (9:1) and screened for Antibacterialactivity between Bita-Sitosterol and Diosgenin. Based on the results, the potent fractions were subjected to isolation of active constituents using column chromatography. Results: The isolated compounds from two fractions n-hexane and Acetic acid were characterized by modern analytical techniques such as UV-visible Spectroscopy, FTIR (Fourier Transform Infrared spectroscopy), NMR Spectroscopy and Mass spectroscopy as Bita-Sitosteol and Diosgenin. Conclusion: The two compounds isolated from potent fractions based on column chromatographywere characterized as Bita-Sitosterol and Diosgenin by modern analytical techniques. Well Diffusion Assay confirm that Bitasitosterol and Diosgeninhave antibacterial activity and can be used as the source for natural bactericides