Application of sesquiterpene lactone in trilobe wedelia in preparation of medicine for resisting tobacco mosaic virus (TMV)
A kind of technology of sesquiterpene lactone, trilobite weed chrysanthemum, applied in application, disinfectant, animal repellent and other directions
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Embodiment 1
[0015] Compound extraction and separation:
[0016] The dried whole plant of Wedelia trilobata was refluxed with 95% methanol and extracted three times (4h, 4h, 3h). Alcohol was extracted to obtain four fractions of petroleum ether, chloroform, ethyl acetate and n-butanol. The chloroform fraction was separated by column chromatography using silica gel (100-200 mesh) and petroleum ether / ethyl acetate system to obtain 5 fractions. Compounds 1 (0.00021%), 2 (0.000061%), 5 (0.0002%) and 6 (0.000052%) were obtained by separating the petroleum ether / ethyl acetate 6:4 fraction through silica gel and Sephadex LH-20 gel. Compounds 3 (0.0002%) and 4 (0.000052%) were obtained by the same separation process of petroleum ether / ethyl acetate 1:1 fraction. The petroleum ether fraction was separated by column chromatography using silica gel (100-200 mesh) and petroleum ether / ethyl acetate system to obtain 5 fractions. The petroleum ether / ethyl acetate 6:4 part was separated by silica gel, ...
Embodiment 2
[0020] 1. Experimental method
[0021] Half-leaf dead spot method of heart leaf tobacco: select healthy and suitable heart leaf tobacco, and place it in a dark room overnight. Select 3 leaves of similar size from each tobacco tree. The test compound is isolated from Wedelia trifida, dissolved in dimethyl sulfoxide (DMSO) into a 10 mg / ml mother solution, stored at 4°C, and diluted with sterile water before use. The required concentration, the final concentration of DMSO 25μl / ml has no effect on the experimental results. One half of each leaf was evenly treated with 100 μl, and the other half was treated with 25 μl / ml DMSO solution. After 3 hours, each leaf was inoculated with 200 μl of 50 μg / ml TMV by rubbing, half of the untreated leaves were used as a positive control, and the leaves that were not treated and rubbed with 25 μl / ml DMSO solution were used as a blank. After 10 minutes, the carborundum on the leaf surface was washed with sterile water. Put it in an insect-free...
Embodiment 3
[0032] 1. Experimental method
[0033] Determination of the activity of phenylalanine ammonia lyase (PAL) in tobacco leaves: Different concentrations of compound 1 were sprayed on the leaves of common tobacco K326, and tobacco leaf samples were taken at 6 hours and 24 hours respectively. Tobacco leaves treated with the same concentration of DMSO served as negative control. Take 0.5g of fresh leaves, add 10% polyvinylpyrrolidone (0.05mol / L, pH 8.8 boric acid buffer preparation) 0.5mL, add 2mL boric acid buffer solution (pH 8.0) containing 5mmol / L mercaptoethanol, grind at 4°C, 10000r Centrifuge for 15 min at 10 min / min, take 1 mL of the supernatant (or dilute it to a suitable concentration), add 1 mL of 0.02 mol / L phenylalanine (0.05 mol / L, prepared in boric acid buffer solution with pH 8.8), add 2 mL of double distilled water, and add 2 mL of double distilled water. For phenylalanine, add 1 mL of double-distilled water to make a total volume of 4 mL. Measure the OD value at a...
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