Abstract

BACKGROUND Plants have evolved abundant defensive secondary metabolites to resist insect herbivores. Lycorine is an alkaloid with insecticidal activity from Amaryllidaceae plants, which the destructive pest Spodoptera litura naturally avoids. Cytochrome P450 enzymes are central to xenobiotic detoxification in insects, but the mechanism by which lycorine acts against S. litura remains unknown. This study aimed to reveal the toxic mechanism of lycorine focusing on P450-mediated detoxification. RESULTS Lycorine exhibited substantial toxicity to first-instar S. litura larvae (LD50 = 0.55 μg larva-1). Subsequently, when fifth-instar larvae were exposed to a sublethal dose (LD30) of lycorine, Lyc disrupted metabolic pathways, damaged Malpighian tubules, and induced oxidative stress. Furthermore, lycorine strongly repressed a CYP6AE gene cluster (CYP6AE47, CYP6AE50, CYP6AE70, CYP6AE138 and CYP6AE139) and decreased total P450 activity to 45% in the Malpighian tubules. RNAi co-silencing of these cluster genes increased larval mortality (+30%) under lycorine treatment. Finally, molecular docking and microscale thermophoresis analyses further confirmed direct binding between Lyc and this CYP6AE gene cluster, with the strongest affinity observed for CYP6AE47 (Kd = 518.5 nM). A key residue, ARG170, may be vital for the interaction between Lyc and CYP6AE47. CONCLUSIONS These results demonstrate that the insecticidal mechanism of Lyc involves suppressing the expression and function of a CYP6AE gene cluster, thereby impairing detoxification capacity, which leads to Lyc accumulation and larval mortality. Elucidation of the detoxification system-targeted mechanism for this plant-derived compound provides a foundation for developing novel, sustainable pest management strategies against S. litura and potentially other noctuid pests. © 2026 Society of Chemical Industry.