This work assesses the antibacterial activity of plumbagin (5-hydroxy-2-methylnaphthalene-l,4-dione) and of methanol, chloroform and aqueous extracts of Plumbago zeylanica L. root against various pathogenic bacteria, and the minimum inhibitory concentrations (MICs). Plumbagin and chloroform extracts of Plumbago zeylanica L. root showed antibacterial activity against Escherichia coli, Salmonella typhi and Staphylococcus aureus. Inhibition against Klebsiella pneumoniae, Serratia marcescens and Bacillus subtilis was moderate, and lower against Proteus vulgaris and Pseudomonas aeruginosa. The methanolic extract exhibited moderate activity and the aqueous extract weak activity against the bacterial strains as assessed by disc diffusion assays. The bioactive compound plumbagin and extract of Plumbago zeylanica root show a wide spectrum of antibacterial activity. The compound shows promise as a new drug for various bacterial infectious diseases.