Application of Lanosta-7,9(11),24-trien-3-one 15,26-dihydroxy-sterane triterpene in preparation of medicament for preventing and/or treating EV71 infection
A technology of ganoderma acid and dihydroxyl, used in ganoderma 7,9(11),24-trien-3-one 15,26-dihydroxy-stane triterpene in the preparation of drugs for preventing and/or treating EV71 infection It can solve the problems of unmentioned and application in the prevention and treatment of enterovirus type 71, and achieve the effect of safe and effective toxic and side effects, alleviating the pain of treatment and improving the quality of treatment.
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Embodiment 1
[0021] Example 1: Toxicity test of Ganoderma acid Lanosta-7,9(11),24-trien-3-one15,26-dihydroxy on host cells (MTT method)
[0022] After 24-48 hours of culture, when the RD cells are almost covered with a single layer, the culture medium is discarded, digested with trypsin, and transferred to a 96-well sterile cell culture plate, 100 μl per well. Place it in a cell culture incubator for 18-24 hours to make the cells grow into a single layer for later use. Dilute the Lanosta-7,9(11),24-trien-3-one15,26-dihydroxy stock solution with cell culture medium and configure five concentration gradients, then add different concentrations of drugs and discard the supernatant 100 μl per well, repeat 3 wells for each concentration, and set up cell control wells (without adding drugs, only culture medium), and then add 100 μl of cell culture medium to each well, place at 37 ° C, cell culture In the chamber, after culturing for 48 hours, add 20ul of MTT solution (5mg / ml prepared in PBS, pH=...
Embodiment 2
[0026] Example 2: Determination of half infection dose (TCID50) of cells by EV71
[0027] Transfer the RD cells cultured into a single layer to a 96-well cell culture plate, and place them in a cell culture incubator for 18-24 hours. The virus liquid was serially diluted 10 times with the maintenance solution (10 -1 …10 -8 ). Discard the culture medium in each well of RD cultured into a monolayer, wash each well with PBS 3 times, add 100 μl of virus solution of different concentrations to each well, absorb at 37°C for 1.5 h, discard the virus dilution solution, and add 100 μl to each well Maintenance solution, 10 repetitions for each concentration, set normal cell control wells. The cytopathic changes (CPE) in each well were observed daily for 3 consecutive days, and the CPE conditions were recorded. The titer of the virus was then calculated by the following formula.
[0028] PD / [log(dilutionbove50%)-log(dilutionbelow50%)]=[(%nextbove50%)-50%] / [(%nextabove50%)-(%nextbelo...
Embodiment 3
[0030] Example 3: Experiment of the antiviral effect of Ganoderma acid Lanosta-7,9(11),24-trien-3-one15,26-dihydroxy on EV71
[0031] 1) Preventive effect of Ganoderma acid Lanosta-7,9(11),24-trien-3-one15,26-dihydroxy on EV71 virus infection
[0032] After 24-48 hours of culture, when the RD cells are almost covered with a single layer, the culture medium is discarded, digested with trypsin, and transferred to a 96-well sterile cell culture plate, 100 μl per well. Place it in a cell culture incubator for 18-24 hours to make the cells grow into a single layer for later use. The drug was diluted step by step with cell culture medium. After dilution, add different concentrations of drugs into the cell culture wells where the supernatant was discarded, 100 μl per well, repeat 3 wells for each concentration, and set up cell control wells (no virus, no drug, only culture solution) and Virus control wells (no drug, virus, culture medium). After drug incubation for 1 hour, discard...
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