Bioactive Constituents Isolated From Orixa Japonica And Toddalia Asiatica

Twenty-three essential oils derived from Chinese Rutaceae plants were screened for insecticidal activity against Liposcelis bostrychophila.Two Chinese medicinal herbs Orixa japonica and Toddalia asiatica were chosen for identification of their bioactive constituents against pests and nematodes.And the possible mechanism of an active compound against Ostrinia furnacalis was also determined.Essential oils of O.japonica and T.asiatica showed contact activity against Blattella germanica(LC50=320.86 and 876.20 ?g/adult,respectively),L.bostrychophila(LC50=131.29 and 339.46 ?g/cm2,respectively),Sitophilus Zeamais(LC50=62.00 and 56.72 ?g/adult,respectively)and Tribolium castaneum(LC50=43.79 and 67.81 ?g/adult,respectively).The essential oil of T.asiatica also showed fumigant activity against L.bostrychophila,S.Zeamais and T.castaneum with LD50 values of 7.37,51.38 and 33.15 mg/L,respectively.Moreover,the two essential oils were highly repellent against L.bostrychophila with percentage repellencies of 89.40%and 97.18%,respectively,at the concentration of 17.0 nL/cm2.The two essential oils possessed nematicidal activity against Bursaphelenchus xylophilus(LC50=127.96 and 475.04 ?g/mL,respectively)and Meloidogyne incongnita(LC50=31.37 and 115.28?g/mL,respectively),as well as larvicidal activity against the fourth instar larvae of Aedes albopictus(LC50=75.20 and 69.06 ?g/mL,respectively),Ae.aegypti(LC50=93.68 and 75.71?g/mL,respectively),Anopheles sinensis(LC50=10.82and11.92?g/mL,respectively)and Culex pipiens pallens(LC50=59.49 and 42.86?g/mL,respectively).On basis of bioassay-guided fractionation,P-caryophyllene(1),caryophyllene oxide(2),?-cadinol(3),a-asarone(4),D-lemonene(5),geraniol(6)and isopimpinellin(7)were identified from the two essential oils.The seven active compounds were evaluated for insecticidal and repellent activity against the three grain storage insects and larvacidal activity against the four species of mosquitoes.They were also evaluated for nematicidal activity against the two species of nematodes.Of the seven isolates,compound 6 was the most active compound in the contact tests(170.41?g/cm2 against L.bostrychophila,18.62 ?g/adult against S.Zeamais and 30.27 ?g/adult against T.castaneum).Compound 6 also possessed the highest fumigant activity against the three stored pests with LC50 values of 1?13 mg/L.Besides,compounds 1,2,5 and 6 were highly repellent against the stored pests with their 4 h percentage repellencies over 80%.In addition,compound 3 and 5 exhibited high nematicidal and larvicidal activity with LC50 values of 15?150 ?g/mL against the two species of nematodes,and 1-23?g/mL against the four species of mosquitoes.Based on bioassay-guided fractionation,a total of 68 active compounds were isolated and identified from the ethanol extracts of the two plants,including 8 new compounds and a new natural product.Moreover,19 compounds were first isolated from O.japonica while 9 compounds were first isolated from T.asiatica.The isolates were tested for their larvicidal activities against the fourth instar larvae of Ae.aegypti,An.sinensis and C.pipiens pallens,as well as nematicidal activities against B.xylophilus and M.incongnita.Bioactive compounds a-terthienyl(8)and ascaridole(11)were isolated from O.japonica for the first time,to the best of our knowledge.a-Terthienyl(8)was the most active compound with its LC50 values of 0.02?0.4 ?g/mL against the three species of mosquitoes,and 0.4?2.0?g/mL against the two species of nematodes.Ascaridole(11)also possessed excellent activity with its LC50 values of 8?46 ?g/mL against mosquitoes,and 29?82 ?g/mL against nematodes.Lunidonine(22)was chosen for further studies.The compound exhibited antifeedant activity against the fourth instar larvae of O.furnacalis with an antifeeding rate of 70.31%at the concentration of 5.0 mg/g.Besides,lunidonine significantly reduced the relative growth rate and the efficiency of conversion of ingested food into biomass of O.furnacalis.After being fed with lunidonine-treated diets,the content of protein in the larval midguts decreased heavily.Moreover,activities of non-specific proteases and a-amylase reduced while activities of GSTs and CYP 450 were activated significantly.Clear evidence of post-ingestive activity of lunidonine to midgut tissues and cells were observed under microscope.In this study,we evaluated the insecticidal and nematicidal activity of the essential oils extracted from O.japonica and T.asiatica,and the bioactive constituents isolated from the ethanol extracts of the two plants using the method of bioassay-guided fractionation,for the first time,to the best of our knowledge.We also investigated the action mechanism of lunidonine against the fourth instar larvae of O.furnacalis for the first time.However,further researches are still needed to investigate their safety issues to humans and non-target organisms and to improve stability of these chemicals so as to develop potential natural pesticides or repellents.In conclusion,both O.japonica and T.asiatica,as well as their active constituents are promise to be developed into eco-friendly pesticides or repellents for control of pests and nematodes.