AbstractIn this study, the inhibition of molting fluid enzymes from Helicoverpa armigera by an aspartic protease inhibitor, Alkalophilic Thermophilic Bacillus Inhibitor (ATBI) purified from Bacillus sp. is reported. The in vitro experiments showed 80% inhibition (IC50= 48 µM) of hemoglobin hydrolyzing and 95% inhibition (IC50= 35 µM) of chitin hydrolyzing activity from molting fluid by ATBI (IC50 value is the ATBI concentration for 50% inhibition of total enzymatic activity). The treatment of H. armigera larvae with 400 µM ATBI recorded 20% larval mortality, 27.77% deformed pupae and 12.22% deformed adults. The LC50 value (Concentration of ATBI calculated to give 50% mortality) calculated for insect population was found to be 330.06 µM. Similarly, significant variations in mean larval and pupal weight, no. of eggs laid per female and percent hatching of eggs were observed at higher concentrations of ATBI. The results may provide the basis for the selection of non‐host inhibitors to develop a H. armigera insecticide formulation.