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Azadirachtin exhibits excellent bioactivities against several hundred arthropods. However, current knowlege of its biochemical effect on B. dorsalis larvae is not deep enough. In this study, integrated LC-MS and GC-MS-based untargeted metabolomics were used to analyze the changes of endogenous metabolites and the biochemical effects of azadirachtin on B. dorsalis larvae. Azadirachtin has excellent bioactivities against B. dorsalis larvae in this study, leading to a longer developmental duration, lower survival rate, and low pupa weight.