Imidacloprid is a neonicotinoid compound, which is a neuro-active insecticide. The over use of this pesticide requires higher awareness about this pesticide. Present investigation was carried out to analyze the effect of imidacloprid on metabolic enzymes amylase, protease and phosphatase in soil isolates Escherichia coli and Brevundimonas Sp. MJ 15 Study on effect of exposure of 10-7 to 10–3 Molar concentrations of imidacloprid for a period of 24, 48, 72 and 96 hrs on three antioxidant enzymes Catalase Peroxidase and superoxide dismutase showed that there was an increase in the activity of all the three enzymes significant increase was observed for doses (10-3-10-7) of imidacloprid. The enzyme activity increased with an increase in the concentration of insecticide at lower doses proving that the inhibitory effect is dose dependent. The present investigation indicates that imidacloprid induces the significant increase in activity of antioxidant enzymes in the soil isolates. This proves that imidacloprid induces oxidative stress in bacterial isolates.