Application of Corylifol A in preparing anti-radiation drug

[0028] Example 1 Extraction of Corylifol A and its characterization

[0029] Corylifol A extraction

[0030] 15kg of psoralen fruit, crushed, immersed and extracted 3 times with 75% ethanol at room temperature for 24 to 36 hours each time, and concentrated under reduced pressure to obtain 5L of extract. The extract is applied to silica gel column chromatography with a volume ratio of 100:1 Gradient elution of petroleum ether-ethyl acetate system to 0:100 to obtain 259g of extract at 5:5; the above extract was subjected to silica gel column chromatography to dichloromethane with a volume ratio of 95:5 to 8:2. Ethyl acetate system gradient elution, thin-layer chromatography detection, collect 5g of Corylifol A-containing fractions, and then preparative liquid chromatography, eluted with a methanol-water system with a volume ratio of 75:25 to 95:5 to obtain Corylifol A pure product 1.6g.

[0031] Characterization of Corylifol A

[0032] Light yellow powder, m/z: 391.1896[M+H] by electrospray ionization mass spectrum signal + , Calculate its molecular formula as C 25 H 26 O 4; 1 H NMR spectrum data are as follows: δ8.11 (1H, s, H-2), 8.07 (1H, d, J = 8.8, H-5), 7.00 (1H, dd, J = 8.8, 2.0, H-6 ), 6.89(1H,d,J=2.0,H-8), 7.38(1H,d,J=2.0,H-2'), 6.88(1H,d,J=8.0,H-5'), 7.30 (1H,dd,J=8.0,2.0,H-6'), 3.40(2H,d,J=7.2,H-1”), 5.43(1H,t-like,J=7.2,H-2”) , 2.06(2H,m,H-4”), 2.11(2H,m,H-5”), 5.14(1H,t-like,J=6.8,H-6”), 1.59(3H,s,H -8”), 1.62 (3H, s, H-9”), 1.76 (3H, s, H-10”). From the above characterization, the chemical structural formula of corylifol A can be determined as follows:

[0033]

[0034] Example 2 In vitro cytotoxicity test of Corylifol A

[0035] 2.1 Cell culture

[0036] HBL-100 cells (ATCC) in RPMI-1640 medium (containing 10% fetal bovine serum, 100U/mL penicillin and 0.1mg/mL streptomycin), MCF-7 cells (ATCC) in DMEM/High Gluose medium (Containing 10% fetal bovine serum, 100U/mL penicillin and 0.1mg/mL streptomycin) at 37℃, 5% CO 2 , Cultivate in an incubator with saturated humidity. When the cells grow adherently to a density of 70-80%, pass them at a ratio of 1/2. When passaging, digest them with 0.25% trypsin in the incubator for 1 to 2 minutes. Count cells that are in good growth phase for the experiment.

[0037] 2.2 Cytotoxicity test

[0038] The CCK-8 method was used to test the effect of the tested drug on the survival rate of HBL-100 and MCF-7 cells.

[0039] (1) Cell inoculation: Take cells in good logarithmic growth phase, digest them with 0.25% trypsin, and prepare a cell suspension at 7×10 3 Cells/well, 100 μL of cell suspension was evenly inoculated in a 96-well plate (filled with sterile PBS around the experimental wells), and cultured in an incubator.

[0040] (2) Drug treatment: After inoculating for 12 hours and waiting for the cells to adhere to the wall, take out the 96-well plate, and add 100 μL of corylifol A (6.25, 12.5, 25, 50, 100 μmol/L) solution (all containing 10% fetal cattle) with corresponding concentrations. Serum medium preparation), continue to cultivate in the incubator.

[0041] Drug treatment plan: The experiment was divided into experimental group (medium containing cells, drugs, irradiation); control group (medium containing cells, irradiation); blank group (medium containing cells, no irradiation).

[0042] (3) Color development: After adding the medicine for 48 hours, add 100 μL of medium solution containing 10% CCK-8 to each well, and incubate in an incubator for 3 hours.

[0043] (4) Detection: Use a microplate reader to measure the OD value at a wavelength of 450nm.

[0044] (5) Calculation: cell survival rate = (OD test group -OD Blank group )/(OD Control group -OD Blank group )×100%.

[0045] 2.3 Experimental results

[0046] Corylifol A's in vitro cytotoxicity test results are as follows figure 1 and figure 2 Shown; from figure 1 and figure 2 It can be seen that Corylifol A has a certain cytotoxic activity against HBL-100 and MCF-7 at a concentration of 50 μmol/L, but has no obvious cytotoxic activity at a concentration of 25 μmol/L or less.

[0047] Example 3 Corylifol A cell radiation resistance experiment

[0048] 3.1 Cell radiation resistance experiment

[0049] Under the condition that the cell survival rate of corylifol A group and the control group was not significantly different under 2.2 in Example 2, 25 μmol/L was selected to test its radiation damage repair effect on HBL-100 and MCF-7 cells. The experiment was divided into blank group (no irradiation, no administration), control group (irradiation, no administration), and corylifol A group (experimental group, irradiation, administration).

[0050] Take the cells in good logarithmic growth phase cultured under 2.1 of Example 2 and digested with 0.25% trypsin to prepare a cell suspension. 3 Cells/well, 100μL of cell suspension was evenly inoculated in a 96-well plate (filled with sterile PBS around the experimental wells); after culturing in an incubator for 12 hours, take out the 96-well plate and add 100μL of the corresponding concentration of drug solution ( All are prepared with a medium containing 10% fetal bovine serum); after continuing to culture for 24 hours in an incubator, the control group and the experimental group are given at the same time 137 Csγ-rays are irradiated once, the radiation dose rate is 1.00Gy/min, and the total radiation dose is 8Gy; 24h after irradiation, the color is developed by CCK-8 method; after 3h, the OD value is measured at 450nm wavelength of the microplate reader. The calculation method of cell survival rate is the same as 2.2.

[0051] 3.2 Data processing

[0052] GraphPad Prism 5 software was used to map and analyze the data. The experimental group and the control group were compared with the data for significant difference test, P <0.05 considered the difference to be statistically significant.

[0053] 3.3 Experimental results

[0054] Corylifol A's cell radiation resistance test results are as follows image 3 and Figure 4 Shown; from image 3 and Figure 4 It can be seen that Corylifol A can repair the radiation damage of HBL-100 and MCF-7 cells at a concentration of 25μmol/L.