Screening And Identification Of Antifungal Plants And The Active Components

Fungicides made a significant contribution to the development of agricultureand food supplies as an important agricultural input material. However, concernsabout the environmental and agricultural safety has been sparked due to theexcessive use of chemical fungicides. Therefore, research and development on lowtoxicity, high efficiency and environmentally friendly new biological fungicides hasbecome imminent. China abounded in resources of medicinal plants, which can beapplied to screen the plants possessing inhibitory effect on plant pathogenic fungi.Under the bioassay-guided fractionation method, the main antifungal componentswere separated, purified and structurally identified as the lead compounds ofbotanical fungicides development. In this study, acetone extracts of 28 kinds ofmedicinal plants were applied to screen the ones with a better inhibition rate of plantpathogenic fungi, of which Polygonum cuspidatum possessed a higher antifungalactivity but less research. So, separation, purification and identification of itsantifungal ingredients were carried out under the bioassay-guided fractionationmethod. The results are as follows:1. 28 kinds of traditional herbal plants were selected with effects ofheat-clearing and detoxification, rheumatism alleviating and insecticidal activities.Then they were further extracted with acetone which possessed a small toxic effectagainst plant pathogens. The antifungal activities of the acetone extracts of 28 herbswere measured with mycelial growth method, with Pythium aphanidermatum,Phytophthora capsici, Fusarium oxysporium, Fusarium solani, Botrytis cinerea andFusarium graminearum as test strains, and the acetone extracts of Eugeniacaryophyllata possessed 100% inhibition rate against all the six test fungi, whileArmeniaca mume, Stemona sessilifolia and Aucklandia lappa possessed 70%inhibition rate against five plant pathogens. The medicinal plants with a betterantifungal activities were further studied with spore germination method, and foundthat the inhibition rates against Botrytis cinerea of these medicinal plants were allabove 90%.2. The antifungal activities of Polygonum cuspidatum ethanol extracts againstthe six test fungi were measured by mycelial growth method, and a certain of antifungal activities were showed at the concentration of 1g/L of the crude extractsagainst the six plant pathogens, of which the inhibition rates against P.aphanidermatum and P. capsici were much higher, with 40.78% and 40.65%respectively. The EC₅₀ experiments showed that the ethanol extracts of Polygonumcuspidatum possessed a highest antifungal activity against P. capsici with EC₅₀ of1.18 g/L, followed by a higher activity against F. solani with EC₅₀ of 1.34 g/L. Theethanol extracts of Polygonum cuspidatum were further extracted by petrolieumether, ethyl acetate and N-butanol, and the antifungal activities were measuredrespectively. The results showed that the ethyl acetate extracts possessed the highestantifungal activity against the six plant pathogens, and all the extracts possessedhigher antifungal activities than ethanol extracts, which indicated that the activeantifungal ingredients mainly exist in the ethyl acetate extracts.3. Under the bioassay-guided fractionation method, we obtained two activeantifungal substances, namely Red-5-a and Red-6-d after purification steps of TLC,silica gel column chromatography and Sephadex LH-20 Sephadex columnchromatography of the ethyl acetate extracts of Polygonum cuspidatum. The EC₅₀ ofRed-5-a against P. capsici and F. oxysporium was 0.17g/L and 7.99g/L respectively,while Red-6-d possessed the EC₅₀ value of 0.009g/L and 0.013g/L against P. capsiciand F. oxysporium respectively. REd-5-a were an needled orange crystals, andalmost insoluble in water, but soluble in alcohol and alkaline solutions. REd-6-dwere white crystals, insoluble in water but soluble in ethanol, acetone and otherorganic solvents. We further determined the two substances by UV, IR,¹H-NMR,¹³C-NMR and other spectroscopic methods, and data comparisons were carried out.Finally, REd-5-a was determined as 1,3,8 hydroxy-6-methyl anthraquinone, or thename of emodin, and REd-6-d was 3,4,5-hydroxy stilbene, or the name ofresveratrol.