The essential peel oil of Citrus limetta var. Mitha (Sweet lime) extracted by steam distillation was assessed for chemical constituents and antimicrobial activity. Gas chromatographic analysis identified 17 constituents among which Limonene (95.98 %) was found as major component followed by camphene (1.79 %), while the remaining terpenes were less than 1%. The results of antimicrobial activity of essential oil tested by disc diffusion method, against different against bacteria and fungi showed that it exhibited maximum zone of inhibition against Bacillus cereus (31.0mm) and Bacillus subtilis (29mm) followed by Staphylococcus aureus (25.3mm), whereas the minimum zone of inhibition was shown by Fusarium oxysporum (11mm) after 48 hours of incubation at their respective temperature (370C for bacteria and 250C for fungi). The inhibition zones, measured after 48 hours and 96 hours, showed that it was active against all tested bacteria and fungi. The results of our study showed that essential oil of Citrus limetta var. Mitha peel can be an effective medicine against different pathogenic microbes.