C21 steroid compound in marsdenia tenacissima and preparation method and application thereof
A technology of compounds and steroids, applied to the preparation of steroids and steroids, chemical instruments and methods, etc.
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Embodiment 1
[0051] The preparation of embodiment 1 Caulis officinalis active extract
[0052] Take 15 kg of dried Tengguan vine, extract it 3 times with 75% ethanol under reflux, the ratio of solid to liquid is 1:8, each time for 2 hours, recover the solvent under reduced pressure, and concentrate to 10L. Extracted three times with equal volumes of ethyl acetate and n-butanol respectively to obtain 115 g of ethyl acetate extract and 173 g of n-butanol extract.
[0053] Among them, the n-butanol layer is rich in C 21 Steroidal compounds, after content determination, its C 21 Steroid content reaches 82%.
Embodiment 2
[0054] Embodiment 2 separates C from the vine of customs clearance 21 Steroids.
[0055] Get the ethyl acetate layer extract 110g of gained in embodiment 1, adopt silica gel open column chromatography to separate, and mobile phase selects sherwood oil: ethyl acetate (100:1-2:1) and dichloromethane: methanol (100: 5-2:1) gradient elution, the obtained fractions were analyzed by silica gel thin-layer chromatography, and 10 eluates were obtained after combining the same fractions.
[0056] Petroleum ether: ethyl acetate = 100:30 fraction 11.0g was separated, using silica gel column chromatography, petroleum ether: ethyl acetate gradient elution, at 100:40, the yellow solid was separated by SephadexLH-20, and then The compounds 1-3 and 5 were obtained by separation by HPLC and recrystallization.
[0057] Petroleum ether: ethyl acetate = 100:40 fraction 8.9g was separated, using silica gel column chromatography, dichloromethane: methanol gradient elution, at 100:10, the solid par...
Embodiment 3
[0061] Example 3 Determination of the proliferative activity of 11 compounds on MC3T3-E1 osteoblast cell line by MTT detection.
[0062] Experimental operation
[0063] (1) Cell culture
[0064] MC3T3-E1 cell line was cultured in 10% fetal bovine serum, 100 μg·mL -1 Penicillin, 100 μg·mL -1 In the DMEM medium of streptomycin, the cell culture environment was 37°C, 5% CO 2 The constant temperature incubator was used to incubate and grow, digested with 0.25% trypsin, and passaged every other day.
[0065] (2) Sample preparation
[0066] Take the ethyl acetate extract of the ethanol extract of Tengguan vine and the isolated monomer compound, weigh an appropriate amount of sample and dissolve it in DMSO solution to prepare a concentration of 100 μmol L -1 The solution was stored in a refrigerator at 4°C for later use.
[0067] (3) Experimental process and grouping
[0068] Take the MC3T3-El cells in good growth state, after digestion, dilute with DMEM medium containing 10% ...
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