关键词: 棉铃虫, 海藻糖, 海藻糖酶, 抑制剂, 栗精碱, Lineweaver-Burk法

Abstract: 【Aim】 The objective of the present study is to investigate the in vitro and in vivo  inhibitory effects of castanospermine, a plant-derived chemical, on the soluble trehalase 1 (Harmtre1) from midgut tissues of the 5th instar larvae of Helicoverpa armigera  and the inhibition mechanism. 【Methods】 Crude Harmtre1 protein in the midgut tissues of the 5th instar larvae of  H. armigera  was isolated and purified by ion exchange chromatography and hydrophobic chromatography. Molecular weight of the obtained Harmtre1 protein was determined by gel filtration chromatography. Different concentrations of castanospermine were employed to determine the in vivo and in vitro inhibition rate and median inhibitory concentration (IC₅₀) on trehalase. The type of inhibition was determined with Lineweaver-Burk’s and Dixon’s mapping methods. 【Results】 The specific activity of enzyme, recovery rate, purification factor and the molecular weight of Harmtre1 was 2.022 U/mg, 67.57%, 23.84 and 67 kDa, respectively. The in vitro inhibition assay showed that the inhibition rates of Harmtre1 by 3.75, 7.5, 15 and 30 μmol/L of castanospermine were 42.00%, 50.30%, 58.32% and 67.33%, respectively. And the median inhibitory concentration (IC₅₀) was 7.32 μmol/L. According to the results of Lineweaver-Burk’s and Dixon’s mapping methods, castanospermine was an effective competitive inhibitor of Harmtre1 with an inhibitory constant (Ki) of 4.9 μmol/L. The in vivo inhibition assay indicated that the Harmtrel activity was markedly inhibited by 30, 15 and 7.5 μmol/L of castanospermine at 10 and 20 h after injection (P<0.05). Correspondingly, the content of lymphal Harmtre1 increased constantly with time after injection. Nevertheless, the concentration of lymphal glucose decreased significantly and the energy supply of H. armigera became disordered at 20 h after injection of castanospermine. 【Conclusion】 Castanospermine is an effective inhibitor of Harmtre1. Our results will supply a theoretical support to the development of effective insecticides in control of H. armigera in the future.

Key words: Helicoverpa armigera, trehalose, trehalase, inhibitor, castanospermine, Lineweaver-Burk method