Imidacloprid, 1-[(6-chloro-3-pyridinyl) methyl]-N-nitro-2-imidazolidinimine, is a neonicotinoid. The present investigation was under taken to evaluate the effect of imidacloprid treatment on amylase and protease activity in soil isolate Escherichia coli (SP-02). The soil isolate Escherichia coli were isolated after enrichment cultures, as imidacloprid tolerant bacteria. The isolate was exposed to imidacloprid of varying concentrations ranging from 10-7 M to 10-3 M for a period of 96 hrs. Treatment with higher dose (10-4 and 10-3 M) of imidacloprid resulted into no significant increase in the activity of amylase and protease enzymes whereas, significant increase was observed in the lower dose (10-7, 10-6 and 10-5 M) of imidacloprid. Present investigation revealed that imidacloprid intoxication suppresses amylase and protease at higher doses of imidacloprid. Reduction in dosage of imdacloprid resulted in increase in the amylase and protease activity. The result indicates that imidacloprid toxicity results in reduced activity of essential metabolic enzymes thus effecting the growth of the Escherichia coli.