ABSTRACT: 1-deoxynojirimycin (DNJ) is a natural D-glucose analogue and has a strong physiological activity in inhibiting ?-glucosidase in vivo. The antidiabetic effects of DNJ in mice or other mammals were extensively explored, but the physiological and toxic roles of DNJ in insects was seldom reported. In this study, the biological effects of DNJ were examined in midgut extracts of fourth-instar larvae of Eri silkworm (Samia cynthia ricini, Saturniidae). Based on nuclear magnetic resonance (NMR) metabonomics technology, we analyzed the alterations of glycometabolism, lipids, and energy metabolism pathways in the midgut of S. cynthia ricini caused by DNJ. Pattern recognition analysis (partial least square-discriminant analysis, PLS-DA) showed that four groups of latex, 0.25% DNJ, 0.5% DNJ and the mixture of 0.5% DNJ and latex (1:1) were distinctly different from the control group. Moreover, several metabolic pathways of DNJ-mediated modulation in the midgut were identified. Compared with the control group, alanine, succinate, glutamate, and fumarate concentrations decreased in three groups of 0.5% DNJ, latex, and the mixture, choline levels increased in two DNJ groups, and trehalose levels increased in all experimental groups. Therefore, these results suggest that DNJ modulated lipid metabolism by limiting the hydrolysis pathways of phospholipids metabolism. Additionally, DNJ has a potent negative effect on energy metabolism by inhibiting the hydrolysis of trehalose, glycolysis and the tricarboxylic acid (TCA) cycle. Overall, DNJ, as a single-ingredient, is an efficient substance for modulating lipid metabolism and inhibiting energy metabolism.