Development of resistance to transgenic crops expressing the Cry toxin from Bacterium thuringiensis (Bt) has been the major concern for the long-term success of Bt crops. Alterations in nonbinding site proteinases and Bt toxin receptors are the two types of mechanisms responsible for Bt resistance in resistant insects. However, little is known about the relative contributions of the two types of mechanisms in the early and late phases of the development of Bt resistance. To address the relative contributions of four nonbinding site proteinases including esterase, total protease, chymotrypsin, and glutathione S-transferase in the early and late phases of the development of Cry1Ac resistance, we analyzed the relationships between nonbinding site proteinases and resistance of three groups of Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) strains with different resistance levels because of different geographic origins and selection pressures. Positive correlation (esterase, glutathione-S-tranferases [GST], and chymotrypsin) and negative correlation (total midgut protease) were observed within the low to moderate group II resistant strains. Such correlations were less obvious within the low to moderate group III resistant strains because of only threefold differences in LC50 values. Relative to the unselected susceptible 96S strain, the two highly resistant group I resistant strains BtI and BtR have the same amounts of esterase, GST, and chymotrypsin and disproportionally decreased the amount of total midgut protease. Overall, the low to moderate resistant strains had the lowest amount of the nonbinding site proteinases. The results obtained suggest that alternations in the nonbinding site proteinases probably can only confer low to moderate levels of resistance and thus are enriched in the early phase of the development of Cry1Ac resistance.