Preliminary Study On Physiological Effect And Insecticidal Mechanism Of Itol A Against Brown Planthopper[Nilaparvata Lugens(St(?)l)]

Itol A,an isoryanodane type diterpenoid compound with novel structure,was isolated from the plant of Itoa orientalis Hemsl.(Flacourtiaceae).The insecticidal activity of itol A had been widely reported,but the insecticidal mechanism was not clear.In this study,the biological activity of itol A agaist rice brown planthopper(BPH)[Nilaparvata lugens(Stal)]was measured in different ways,and the effects of physiological and biochemical responses and proteomics of BPH exposed to itol A were studied.The aim of this paper was to provide theoretical basis for defining the insecticidal mechanism of itol A,and provide scientific basis for the development and utilization of itol A as a new insecticide or pilot structure for the prevention and control of BPH.The results are as follows:The results of rice stem dipping test showed that itol A had significant insecticidal activity against 3th instar nymph of BPH,and the median lethal concentration(LC50)values of 48 and 72 hours were 741.99 mg/L and 438.05 mg/L,respectively.The drip method indicated that itol A had contact toxicity to the macropterous male and female of BPH,and the median lethal dose(LD50)values of 24 hours were 0.18 ?g/insect and 0.58 ?g/insect,respectively,and the symptoms of exercise imbalance and leg fibrillation were found in the poisoning insects.In addition,itol A showed significant antifeedant activity on the macropterous female of BPH through the honeydew method,the median antifeedant concentration(AFC50)values were 248.17 mg/L and 148.57 mg/L after treatment for 48 and 72 hours,respectively.The effects of itol A on the acetylcholinesterase(AChE),Na+/K+-ATPase and Ca2+-ATPase of BPH were separately measured in vivo and in vitro.AChE activity assay showed that itol A had no significant effect on AChE activity both in vivo and in vitro.The activities of Na+/K+-ATPase and Ca2+-ATPase in vivo were both significantly impact caused by itol A,and the activities were decreased with the increasing dose,or increased in a short time then decreased rapidly with the increasing time,and eventually were significantly inhibited.In vitro,the activity of Na+/K+-ATPase was significantly inhibited by itol A at the concentration of 10.00 mg/L,the inhibition rate was 12.30%,nevertheless,the inhibition rate was not increased with the increasing concentration of itol A.However,the effect of itol A on Ca2+-ATPase in vitro was opposite,the activity was significantly increased with 14.10%higher than the control with the concentration raising to 100.00 mg/L.Results showed that,itol A had significant effects on the activities of Na2/K+-ATPase and Ca2+-ATPase in vivo,but it may not directly act on these two ATPases,it may be the post-effect on the body.The effects of itol A on detoxification enzymes and protective enzymes of BPH were studied in vivo.The activities of glutathione S-transferase(GSTs)and mixed-functional oxidase(MFO)were both significantly inhibited,and the inhibitory effect was enhanced with the increasing dose or time.The activity of carboxylesterase(CarE)was complex,and its activity fluctuated with the prolongation of treatment time,showing a tendency of inhibition-return to normal-inhibition-activation-return to normal.The activity of acid phosphatase(ACP)of BPH was activated,and the activity of each treatment was both significantly higher than the control.The activity of alkaline phosphatase(AKP)was not significantly changed.The activities of superoxide dismutase(SOD),catalase(CAT)and peroxidase(POD)were both significantly influenced,and the activities were decreased with the increasing dose,or increased first and then decreased rapidly with the increasing time.The differential proteomics of BPH exposed to itol A was studied by the technology of Tandem mass tag(TMT),LC-MS/MS and combined with the database.There were 35 differentially expressed proteins,of which 28 were down-regulated,and others were up-regulated,and involving multiple functions such as metabolism,redox,signal transduction,transcription,protein processing,RNA processing,structural proteins.The above test results show that itol A had significant insecticidal toxicity,contact toxicity and antifeedant activity against BPH which could effectively interference the main detoxification enzymes,protective enzymes and ATPases as well as a variety of important protein expression,disrupt the normal physiological metabolism,thus itol A showed high insecticidal effect.Therefore,it has high research value and broad development prospect of itol A as a new type of medicine or lead structure for controlling BPH.