Preparation and application of Daphnane type macrocyclic diterpenoid compound
A technology of compounds and polymorphs, applied in the fields of steroids, organic chemistry, drug combinations, etc., can solve the problems of inability to effectively cure colorectal cancer, death of patients, and unsatisfactory treatment effects.
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[0066] The second aspect of the present invention provides a preparation method of the compound provided by the first aspect of the present invention, comprising: preparing from Daphne genkwa Sieb.et Zucc extract.
[0067] The preparation method provided by the present invention may include: providing Genkwa genkwa extract. The genkwa extract is usually the extract of genkwa flower buds, and the extract can be alcohol extract. The method of providing the Daphne genkwa extract should be known to those skilled in the art. For example, for the alcohol extract, it can usually be obtained by leaching with an alcohol solvent, and the product obtained by the leaching can usually be concentrated. The alcoholic solvent can generally be ethanol or the like.
[0068] The preparation method provided by the present invention may also include: providing an extract of Daphne genkwa extract. When providing the extract of Daphne genkwa extract, some less polar solvents are usually used, and ...
Embodiment 1
[0081] Preparation of Daphnane-type macrocyclic diterpenoids:
[0082] 1) Take 15 kg of Daphne genkwa flower buds and grind them into coarse powder. Add 3 times the volume of 95% ethanol, soak for two weeks, filter under reduced pressure, recover ethanol under reduced pressure, and concentrate the remaining liquid to a thick paste with a relative density of 1.20. The step of soaking and extracting was repeated more than three times to finally obtain 2 kg of Daphne genkwa ethanol extract.
[0083] 2) The ethanol extract of Daphne genkwa was dissolved in 3L of water, and extracted three times with equal amounts of petroleum ether and ethyl acetate in sequence. The petroleum ether extracts were combined and concentrated under reduced pressure to obtain 400 g of petroleum ether extracts.
[0084] 3) Apply XDA-7 macroporous resin column to preliminary segmentation of petroleum ether extract, and use 60%, 85%, 90%, 95%, 100% methanol-water gradient elution respectively to obtain e...
Embodiment 2
[0117] Cell culture:
[0118] The SW620 cell line used the L-15 medium (Gibco BRL, USA) supplemented with 10% fetal bovine serum and 1% double antibody, and the RKO cell line used the DMEM medium (Gibco BRL, USA) supplemented with 10% fetal bovine serum and 1% double antibody. , USA), the LoVo cell line was cultured in F12k medium (Gibco BRL, USA) supplemented with 10% fetal bovine serum and 1% double antibody, all cultured in 5% CO 2 The saturated humidity is 37 °C in a constant temperature incubator.
[0119] MTT experiment:
[0120] Using the method reported in the literature (Carmichael J, DeGraff WG, Gazdar AF, Minna JD, Mitchell JB. Evaluation of a tetrazolium-based semiautomated colorimetric assay: assessment of chemosensitivity testing. Cancer Res. 1987, 47 (4): 936-942.) , SW620, RKO, LoVo cells were divided into 5×10 3 Inoculate in a 96-well culture plate at a density of 1 / mL, add various concentrations of test samples, and incubate for 48 hours at 37°C, 5% CO2, 9...
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