Inhibition Effects Of Plant Extracts Against Phenoloxidase From Spodoptera Litura Fabricius

Phenoloxidase(PO) is widely distributed in animals, plants, microorganisms and other organisms, which is a key enzyme in melanin synthesis. PO play an important role in metamorphism developing, immune and defense systems, therefore,it can be used as an important target to control the pest insect. It is intresting to screen for a new, effective biological pesticide which targeted against the PO from insect. In this paper, properties of PO from Spodoptera litura Fabricius was studied, and 5 kinds of plant extracts was derived with 6 different solvents to verify their inhibitory effect against PO, among which, water extract of Ailanthus altissima and ethanol extract of Ginkgo biloba L. was selected to assay the toxic effect against S. litura larvae. Detailed results as below: 1. The characteristics of phenoloxidaseProperties of phenoloxidase(PO) from Spodoptera litura Fabricius was determined with catechol as substrates. Data showed that: the optimum p H for PO from S. litura is 6.7~7.0, and the enzyme has the maximum stability at p H 6.8; PO showed the highest activity at 30 ℃, while it has the best thermal stability between 20~35 ℃; when the amount of enzyme was limited to 0.1m L in a total 3 m L assay mixtures, the reaction velocity increased with the concentration of substrate, and when the final concentration of catechol is 10 mmol/L, it reached the highest velocity, then kept constant; the Km value and Vm is 7.980 mmol/L and 204.082 ?A/mg·10s respectively, with catechol as substrate. 2. Inhibition effects of plant materials extracted by parallel extracting method against POInhibition effects of leaf-extracts of Phyllostachys viridis(Young) Mc Clure, Ginkgo biloba L., Sambucus williamsii, Juglans Regia L., Ailanthus altissima and barks-extracts of Ailanthus altissima against PO from S. litura was determined with microplate reader. Data showed that: when the 5 plants(lesf or bark) were treated with 6 different solvents(polarity: deionized water > ethanol > acetone > ethyl acetate > chloroform > petroleum ether), all the extracts showed inhibitory effects against PO, and the inhibition was gradually enhanced along with the increasement of extracts concentration, but the effect of inhibition were diffenent: water extracts of Ailanthus altissima, chloroform extracts of Sambucus williamsii, ethanol extracts of Ginkgo biloba, acetone extracts of Phyllostachys viridis(Young) Mc Clure leafs and water extracts of Ailanthus altissima bark showed the strongest inhibitory effect against PO and the IC50 were 550、500、375、175、800 mg/L respectively; inhibition kinetics study shows that the inhibition effect of 3 kinds of extracts against PO is reversible, they were extracts of Phyllostachys viridis, Ginkgo biloba and Ailanthus altissima. The acetone extract of Phyllostachys viridis is a noncompetitive inhibitor, and the Ki value is 158.2 mg/L. The inhibitory effect of extract from Juglans regia was weaker, not stable with the change of the extract concentration; while the inhibition effect of extracts from Sambucus williamsii against PO is unreversible. 3. Inhibition effects of plant materials extracted by sequential extracting method against POThe inhibition effect of the extracting extracts of Ginkgo biloba and Ailanthus altissima derived with sequential extracting method were stronger than the extracts derived with parallel method, and the inhibition effect of their ethanol extract against PO were greaty enhanced, IC50 were 400 mg/L and 375 mg/L, respectively, the inhibitory effect increased by 20% and 20%. 4. Primary purification of ethanol extract from Ginkgo biloba and water extract from Ailanthus altissimaChloroform extraction of ethanol sequential extract from Ginkgo biloba has the strongest inhibition effect against PO after further purification, and with the increase of the extract concentration, the inhibition effect increased linearly, inhibition effect was 61.03% and 12.31% higher than control when the extract concentration was 200 mg/L; the 90% ethanol sediment of Ailanthus altissima has the best inhibition effect against PO, and Inhibition effect was 57.32% and 49.35% higher than control when the extract concentration was 200 mg/L, data suggests that polysaccharide is the main ingredient which has inhibition effect against PO in the Ailanthus altissima estracts. 5. Bioactivity assay of ethanol extract of Ginkgo biloba and water extract of Ailanthus altissima against the larvases of Spodoptera litura FabriciusThe bioactivity of ethanol extract of Ginkgo biloba and water extract of Ailanthus altissima against the larvases of Spodoptera litura Fabricius were assayed by leaf disc test method. Data showed that: both the two extracts had stomach toxicity and antifeedant activity to the larvaes, antifeedant rates were 77.75% and 89.30% after 2 d, and corrected mortalities were 57.14% and 43.14% after 8 d, when the concentration of extracts were 32 g/L.