A pimarane type diterpenoid compound having an oxygen-containing five-membered ring, a preparation method and application thereof
A technique for a diterpenoid and a five-membered ring, which is used in the preparation of antitumor drugs, in the field of diterpenoids, and achieves the effect of strong inhibitory activity.
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Embodiment 1
[0036] Embodiment 1. Preparation of compounds of the present invention
[0037] 1. Prepare the total crude extract
[0038]1) Pick a small amount of mycelium of the bacterial strain of Eutypella sp.D-1 whose preservation number is CCTCC NO: M 2013144 from the plate, inoculate it into a 250mL Erlenmeyer flask for seed culture, and each Erlenmeyer flask contains 100mL of seed medium, Cultivate at a constant temperature on a shaker at 28°C with a rotation speed of 180r / min, and cultivate for 3.5 days to obtain a first-class seed liquid, insert the same seed medium according to the inoculation amount of 5% (v / v), and cultivate at 28°C and 180r / min on a shaker for 3.5d , to obtain fresh secondary seed liquid for fermentation and cultivation. Then the seed culture liquid is inoculated into the Erlenmeyer flask of 2000mL and carries out expanded cultivation, and the fermentation medium of 400mL of each Erlenmeyer flask, inserts fresh secondary seed liquid according to the inoculum s...
Embodiment 2
[0052] Embodiment 2 The in vitro antitumor activity experiment of the compound of the present invention:
[0053] Method: CCK8 method (Tominaga H, Ishiyama M, Ohseto F, et al.A water-solubleetrazolium salt useful for colorimetric cell viability assay[J].Analytical Communications,1999,36(2):47-50.)
[0054] Human pancreatic cancer cells PANC-1, human breast cancer cells MCF-7, human colon cancer cells HCT-116, human chronic leukemia cells K562, and human pancreatic cancer cells SW1990 in logarithmic growth phase were respectively taken at 5000 cells / well. The cell density was seeded in a 96-well plate and placed in 5% CO at 37°C. 2 cultured in a constant temperature incubator. After 24 hours, add 10 μL of samples of different concentrations to the 96-well plate, so that the final volume of the solution in the plate is 100 μL. After continuing to cultivate in the incubator for 48 hours, add 10 μL of CCK-8 solution and continue to incubate for about 1 hour. Take it out, and use ...
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