Benzyne compound and preparation method thereof, as well as application in control of rice pathogens
A compound and rice disease technology, applied in the field of biological control, can solve the problems of no formal commercial product, unsolved industrial production technology, etc., and achieve the effects of easy industrial production, good control effect, and wide source of raw materials
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[0019] The preparation method of benzyne compound provided by the invention, its steps are:
[0020] 1) Liquid culture and fermentation product processing: Put the flower face mushroom ( Lepista sordida ) (Preliminary report on the inhibitory activity of A. variegata on rice thin strip disease, China Agricultural Science Bulletin, 2012, 28 (12): 202-205.) The slant strains were first inoculated in a Erlenmeyer flask containing 150mL potato liquid medium for activation , the activation conditions are rotation speed 230 r / min, culture temperature 28°C, culture time 7 days, and then carry out large-scale liquid submerged fermentation. The medium formula used is: per liter of water contains 200g of potatoes, 20g of glucose, 5g of peptone, pH Naturally, 0.1Mpa, sterilized at 121°C for 30 minutes, and cultured in a constant temperature shaker at 25-28°C, 180-230 r / min. After 10-15 days of fermentation, the mycelium is separated from the fermentation broth by centrifugation. The...
Embodiment 1
[0028] Use potato glucose medium (containing 200g of potato per liter of water, 20g of glucose, natural pH, 0.1Mpa, sterilized at 121°C for 30min), the activated strain Lepista sordida 3 L of liquid fermentation was cultured on a shaker at 28 °C for 12 days. After bacterial cell filtration, the fermentation broth was concentrated to 0.5 L, the fermentation broth was extracted with ethyl acetate, the ethyl acetate part was dehydrated with anhydrous sodium sulfate, concentrated, then dissolved in methanol and filtered to obtain methanol-soluble crude extract (186.6 mg, brown extract).
[0029] Dissolve 186.6 mg of the methanol crude extract in the previous step with an appropriate amount of methanol, perform reverse-phase silica gel (30 g) column chromatography, and elute with different gradients of methanol-water (containing 1‰ formic acid) at a flow rate of about 15 mL / min, collect 20 mL in each test tube, take samples from each test tube for thin-layer chromatography (deve...
Embodiment 2
[0036] After the blast fungus was cultivated to produce spores, the colony of blast fungus oryzae was washed in a conical flask with sterile water, shaken fully, filtered with sterile gauze to obtain its spore suspension, and the spore concentration was obtained by microscope observation and statistics. 7.5×10 5 individual / mL. The sample was dissolved in methanol and diluted with 0.1% Tween 80 sterile water. Pipette 50 μL of Magnaporthe grisea spore suspension and 50 μL of the diluted sample solution respectively, and drop them on the recess of the concave slide. The contents of the final samples were 1 μg / 100 μL, 2 μg / 100 μL, 4 μg / 100 μL, and 8 μg / 100 μL, respectively. Mix gently and place in a Petri dish lined with wet filter paper. A blank control was set up with methanol, and each treatment was replicated three times, and placed in a 25 ℃ incubator for 12 hours of hanging drop culture. Microscopically observed the morphological changes of spore germination in each trea...
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