Studies On The Separation, Ultrastructure Effect On Mythimna Separata And Agrotis Ypsilon Of Periplaocoside NW

To study and develop the precious insecticidal plant Periploca sepium Bunge,the insecticidal constituents were isolated by bioassay-guided fractionation,and the effect of periplocoside NW on the midgut tissue of Mythimna separata larvae studied by observation of semeiology and laser scanning confocal microscopy,and transmission electron microscopy in this thesis.The main results were as follows:1.By means of non-pressurized column chromatography and preparative HPLC four compounds,G1,G2,G3,G4 were isolated from the methanol extract of Periploca sepium Bunge in the guidance of bioassay-guided fractionation.On the basis of spectral data,G1,G2,G3,G4 were identified as periplocoside NW,periplocoside E,periplocoside A and 2-O-acetyl-β-D-dit-(1→4)-β-D-cym-(1→4)-β-D-cym-(1→4)-β-D-cym-(1→4)- L-oleandronic acid–δ-lactone respectively.2. Periplocoside NW and fluorescent chemical MIA was constructed,and obtained FNW.It was found that FNW emit green fluorescence under 405nm light. The midgut of Mythimna Speparata larvae were respectively incubated in vivo and in vitro with FNW. After turned to frost slice, midgut was observed under laser scanning confocal microscopy. It was showed that midgut holding FNW emits strong green fluorescence.And It can be clearly observed midgut has been damaged and the cell membrane has been disintegrated, which suggested that Periplocoside NW can be combined with binding site of midgut of larvae of Mythimna Speparata.3. By the observation of semeiology, the results indicated that Mythimna Speparata larvae what were fed on Periplocoside NW were not excitement symptoms.Its symptoms appeared after about 4 hours.As time was prolonged,the gastropods have become prominent, but not normal contraction.Final result of abdominal swelling, paralysis flaccid paralysis increased, test insects died. A further tem observation showed that the microvilli layer of goblet cells and columnar cells in the midgut of Mythimna separate larvae were damaged, Thus, its membrane structure was destructed and disappeared, which made the cells disintegrated.PeriplocosideNW also caused the endoplasmic reticulum expanded,vesiculations appeared,ribosomes falling off, mitochondrial swelling, a blank bright area to become fusion, cristae blurred and bilayer to become incomplete. But the intestinal wall cells and organelles of Agrotis ypsilon larvae were not injuried. It showed that the effect of Periplocoside NW on cells and organelle of the midgut of Mythimna separate larvae might be one reason for the death of the tested insects,and it was possible that the midgut cells were the initials target of Periplocoside NW.