| Flaveria bidentis (L.) Kuntze grows strongly, and is an invasive plant in Hebeiprovince of China. The allelochemicals from Flaveria bidentis (L.) Kuntze has a stronginhibition on the growth of the other plants and a good herbicidal activtity. In this study,the chromatographic techniques were used to separate the herbicidal component from theroot of Flaveria bidentis (L.) Kuntze, then the structure of the herbicidal active substancewas analyzed by LC-MS, elemental analysis and1H-NMR. At the same time, theherbicidal mechanism of the component from Flaveria bidentis (L.) Kuntze was tested.1. The herbicidal component from the root of Flaveria bidentis (L.) Kuntae wasseparated via liquid-liquid extraction, TLC, and HPLC. The result of liquid-liquidextraction indicated that petroleum ether was the optimum extraction solvent. Thepetroleum ether extraction was separated by the method of TLC and the purple blue withthe strongest bioactivity was obtained. The purple blue obtained from TLC analysis wasdetected by HPLC and the results showed there were two peaks named as compound I andcompound II with the retention time of4.069min and4.986min respectively. Thecomponent with retention time of4.986min had a higher herbicidal bioactivity, and theinhibitional rate on the growth of D. sanguinalis could was more than90%.2. The structure of the compound II with herbicidal activity was analyzed by LC-MS,elemental analysis and1H-NMR. The results of LC-MS detection showed that the [M+H]+ion of compound II was m/z247.979, the molecular weight of compound II was246.949.The elemental analysis indicated the content of C, H, N, S in compound II was58.165%,3.355%,0.145%and37.280%respectively, and the atomic number of C, H, N and S incompound II was11.97,8.29,0.026and2.88respectively. The elemental of N could beignored, so the molecular formula of compound II was C12H8S3. The NMR spectrumshowed that chemical shifts in1HNMR were δ7.22(dd, J=5.1,1.1Hz,1H),7.18(dd, J=3.6,1.2Hz,1H),7.08(s,1H),7.04–7.00(m,1H). Compound II was determined asα-terthienyl according to the data of1HNMR and the molecular formula. Herbicidalactivity of α-terthienyl was assayed by foliar treatment, the symptoms were monitored at24h post treatment and the result showed that when the concentration of α-terthienyl was50mg·L-1, the degree of inhibitionrate of growth was four, and when100mg·L-1, thedegree of inhibitionrate of growth was five. From these results, we can see that the α-terthienyl has a high herbicidal activity3. High-speed counter-current chromatography (HSCCC), a support-free liquid–liquidpartition chromatographic technique, eliminates the irreversible adsorption of sample ontothe solid support, and has been widely used in the preparative separation of naturalproducts. In this study, the two-phase solvent system was screened by measuring thepartition coefficient and the result showed that the solvent system composed ofn-hexane–ethyl acetate–methanol–water (6:4:5:5, v/v) was optimal for the HSCCCseparation. The methanol extract from the root of F. bidentis (L.) Kuntze was isolated byHSCCC with the conditions that the flow rate was2mL· min-1, the apparatus was rotatedat900r·min-1and the detecting wavelength was214nm. The sample solutions wereseparated as three peaks in the chromatogram, which were numbered sequentially as1,2,and3. The effluents of the three peaks were evaporated under reduced pressure, and theherbicidal activity of the concentrates was determined. The results showed that peak3hadthe strongest herbicidal activity. Therefore, the purity of peak3was detected by HPLC.The results showed that peak3mainly includes two components and it was same as theresult of TLC. But compared with TLC, the HSCCC was easy and rapid, further more itreduce the loss of sample in the exprement.4. The herbicidal mechanism of the component from the root of Flaveria bidentis (L.)Kuntae was tested in this research. The effect on inhibition of seed germination and growthwas determined and the chlorophyll content, the beta-carotene content, the MDA content,superoxid anion radical content, hill activity, water metabolism and defense enzymeactivity were examined. The length of root and stem were stronger inhibited underillumination than under darkness. After treated with herbicidal component, the relativedamage rate of chlorophyll and beta-carotene was higher under illumination than underdarkness. Also, the MDA content, superoxid anion radical content and the relative damagerate of hill activity were higher under illumination than under darkness. The moisture lossrate of D. sanguinalis was growing with the contant growth of herbicidal component. Theeffect of α-terthienyl on the PAL, POD and PPO under illumination was higher than thatunder darkness, simultaneously. These results showed that the herbicidal activity of thecomponent was enhanced under illumination, and it had an effect on photosynthesis, watermetabolism and defense enzyme activity. |
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