The essential oil of the leaves of Psidium guajava grown at Ujjain M.P. (India) was isolated and analyzed by gas chromatography coupled with mass spectrometry (GC-MS). The components of the essential oil were identified by comparing their retention indices and mass spectra fragmentation patterns with those stored on the MScomputer library and also from the published literatures. The present study describes the phytochemical profile and anti-microbial activity of essential oil of P. guajava. Furthermore, anti-microbial activity of oil was evaluated using agar well diffusion method. The anti-microbial test results showed that the oil had a potential anti-microbial activity against all twelve Gram+ve and Gram-ve bacterial strains such as: Staphylococcus aureus, Streptoccocus faecalis, Bacillus subtillis, Lactobacillus spp., Enterococcus aerogenes, Acinetobacter spp.(Gram Positive) and Escherichia coli, Proteus vulgari, Enterobacter aerogenes , Salmonella typhimurium, Pseudomonas aeruginoso, Klebsiella pneumoniae (Gram Nagative) . Essential oil showed maximum zone of inhibition and minimal inhibition concentration against Bacillus subtillus and Escherichia coli bacterial strains. These results permitted the conclusion to be made that, it is the first report of the GCMS analysis and anti-microbial activity on a P. guajava., a naturally growing species from Malva Region of India.