Kojic acid and kojic acid derivative, and preparation method and application thereof
A technology of kojic acid and medicine, applied in the field of kojic acid and kojic acid derivatives and their anti-inflammatory uses, can solve the problems of few reports on the anti-inflammatory activity of kojic acid and its derivatives, etc.
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Embodiment 1
[0033] Fermentation and extraction of embodiment 1 bacterial classification
[0034] Aspergillus versicolor (Aspergillus versicolor), purchased from the American Type Culture Collection (ATCC), the preservation number is ATCC 28286. The strain was revived by streaking, inoculated into PDA solid medium, and activated in a 28°C incubator for 5 days. Use a scalpel to cut out small pieces of colonies from the PDA medium, inoculate them into PDB liquid medium, and cultivate them on a shaker at 28° C. and 160 rpm for 5 days to obtain seed liquid. Then the seed solution was inoculated into rice solid medium, and cultured statically for 30 days in a constant temperature and humidity chamber at 28°C. Take the fermented product of rice, add 400 mL of ethyl acetate for every 200 g of the fermented product, cold-soak and extract twice, combine the extracts and concentrate under reduced pressure until there is no ethyl acetate smell to obtain ethyl acetate extract.
Embodiment 2
[0035] The separation and identification of embodiment 2 trexone A
[0036] The extractant was chromatographed on a macroporous resin column, adsorbed overnight, and eluted with 10%, 30%, 50%, 70%, 90%, 100% methanol-water gradient, and each gradient was combined and spin-dried until there was no liquid to obtain 6 a subcomponent. The fourth subcomponent (70% part) was separated and purified by semi-preparative high performance liquid chromatography, mobile phase: 65% MeOH, flow rate 4mL / min, detection wavelength 210nm, peak time 34.8-35.6min. The peak was picked up and evaporated to complete dryness to obtain 12.5 mg of white solid. The white solid was subjected to high-resolution mass spectrometry and one-dimensional two-dimensional NMR analysis results as follows Figure 1-8 , the structure of the compound is determined to be High resolution mass spectrometry [M-H] – m / z 419.1339, calculated to be 419.1348 (C 21 h 23 o 9 ), the molecular formula of the compound is C...
Embodiment 3
[0039] Example 3 NO release inhibition experiment
[0040] Mouse macrophage cell line RAW 264.7 was cultured in high glucose DMEM medium supplemented with 1% penicillin and streptomycin at 37°C, 5% CO 2 cultivated under conditions. First, the MTT method was used to test the cytotoxicity of the compounds on RAW 264.7 cells. Both compounds had no cytotoxicity, IC 50 >100 μM. RAW 264.7 cells were counted with a hemocytometer, and the cell concentration was adjusted to 5×10 3 cells / well and cultured on a 96-well plate for 24 hours. After that, LPS (1.0 μg / mL) was added, and the compounds were diluted with culture medium to a final concentration of 50, 25, 12.5, 6.25, and 3.125 μM, respectively, and three replicate wells were set up in the sample addition group and the blank group. After continuing to culture for 24 hours, the amount of NO released was tested with a NO kit, and the OD value of each well was measured with a microplate reader at a wavelength of 550 nm. Inhibitio...
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