Cotton plants produce gossypol as a major secondary metabolite to resist insect herbivores and pathogens. Helicoverpa armigera may employ multigene families of detoxification enzymes to deal with this metabolite. So far, the strength of the transcriptional response to gossypol detoxification in the cotton bollworms remains poorly understood. Here, we investigated the genomewide transcriptional changes that occur in cotton bollworm larvae after one generation feeding on various host plants (cotton, corn, soybean and chili) or an artificial diet. Six genes potentially involved in detoxification of xenobiotics were highly upregulated in bollworms fed on cotton, and the expression of five of these differed significantly in insects that fed on gossypol diet compared with the artificial diet. When these six genes were downregulated using RNA interference, downregulation only of CYP4L11, CYP6AB9 and CCE001b led to reduced growth of bollworm larvae feeding on gossypol diets. These data suggest that the three genes are involved in response of H. armigera to gossypol of cotton. Our results proved that H. armigera may have a broad mechanism for gossypol detoxification.