Preparation method as well as structural partial characterization and application of gelidium pacificum okam polysaccharide with immunomodulatory effect
A polysaccharide and immunomodulatory technology of lilyflower, which is applied in the field of natural macromolecules to achieve the effects of promoting immune recovery, inhibiting inflammation and improving human immunity.
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Embodiment 1
[0043] 1. Extraction process
[0044] The dried dashi cauliflower is removed by hand, soaked to remove impurities, dried, crushed, and passed through a 20-mesh sieve. The dashi cauliflower powder is first soaked for 20 minutes, and the material-to-liquid ratio is 1:30. Ultrasound-assisted extraction at 80°C for 2.5 hours to obtain a water extract, 60 °C and concentrated under reduced pressure to obtain a concentrated solution.
[0045] Add twice the volume of absolute ethanol to the concentrated solution, let it stand at room temperature for 12 hours to precipitate polysaccharides, centrifuge at 8000 rpm for 10 minutes, and collect the precipitates at 4°C to obtain crude polysaccharides.
[0046] The crude polysaccharide was dissolved in distilled water, protein was removed by Sevage method, trichloroacetic acid with a final concentration of 3% was added to the concentrated solution, left standing for 12 hours, centrifuged at 8000 rpm for 10 minutes, and the supernatant was ta...
Embodiment 2
[0062] Materials and reagents: Dashi cauliflower polysaccharide A-11 (self-made), DMEM medium (Sigma), LPS (Sigma), fetal bovine serum (Sigma), Griess reagent (Sigma).
[0063] Instrument constant temperature incubator (Shanghai Boxun Industrial Co.), microplate reader (CORONA, Japan), microscope (Shanghai Optical Instrument Factory), 96-well plate (BD, USA).
[0064] 1. The effect of polysaccharides on cell viability
[0065] Cell viability was determined by MTT method. THP-1 cells were divided into six groups with 3 parallels in each group: a blank group was added with 200 μL RPMI-1640 culture solution; a positive control group was added with RPMI-1640 culture solution containing 2 μg / mL LPS; four test groups were added with 2 μg / mL LPS mL LPS and RPMI-1640 culture solution with different concentrations of polysaccharides (1, 5, 10 and 25 μg / mL); cultivated under these conditions for 24 hours. Centrifuge at 1000r / min for 5 minutes to collect cells, add 20 μL of MTT solutio...
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