Application for sulfated mannose glucuronic acid hexasaccharide in prevention and treatment of atherosclerosis
A technology for atherosclerosis and hexasaccharide of uronic acid, which is applied in the directions of medical preparations containing active ingredients, non-central analgesics, cardiovascular system diseases, etc. and other problems, to achieve the effect of inhibiting absorption effect and small toxic and side effects
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Embodiment 1
[0056] The preparation of embodiment 1 sulfated mannoglucuronate hexasaccharide
[0057] 1. Preparation of mannoglucuronide oligosaccharides
[0058] Extract the dried kelp with 30 times the mass of distilled water at 100°C for 3 hours, filter the extract, concentrate, add ethanol to a final concentration of 75% to precipitate, let it stand for 12 hours, collect the precipitate, sink to the point of vacuum drying to obtain laminarin . Dissolve the laminarin sample in a sulfuric acid solution with a mass concentration of 4% (the ratio of solid to liquid is 60mg / mL), heat and reflux for 5 hours, neutralize with barium hydroxide to PH=6-7, centrifuge, and concentrate the supernatant to the original One-fifth of the volume, the concentrated solution is subjected to active carbon column chromatography, firstly equilibrated with distilled water, and then eluted with a gradient of 50%-90% ethanol, and the 50%-90% ethanol eluate is concentrated to one-fifth of the original volume 1....
Embodiment 2
[0065] Example 2 Regulation of Sulfated Mannoglucuronate Hexaose on Inflammatory Factors
[0066] THP-1 cells were seeded in 6-well plates (4 × 10 5 Cells / well, RPMI-1640 complete culture solution containing 10% FBS), phorbol ester (PMA) was added to a final concentration of 100ng / ml, and culture was continued for 48h to induce monocytes to differentiate into macrophages. The experimental group was added with 0.25mg / ml, 0.5mg / ml, 1mg / ml sulfated mannoglucuronide hexaose respectively. Cultivate for 48 hours, collect the culture solution, and remove the precipitate at 4000 rpm for 5 minutes. ELISA kits were used to detect the release levels of TNF-α and IL-1β.
[0067] The result is as figure 1 As shown, the sulfated mannoglucuronate hexaose with a concentration of 0.5 mg / ml can significantly reduce the inflammatory factor TNF-α ( figure 1 A) and IL-1β ( figure 1 The expression level of B) increased to 1mg / ml, and the effect of reducing the expression level of inflammatory ...
Embodiment 3
[0068] Example 3 Inhibitory regulation of sulfated mannoglucuronide hexaose on NF-κB signaling pathway
[0069] THP-1 cells were seeded in 60mm culture dishes (2×10 6Cells / well, RPMI-1640 complete culture solution containing 10% FBS), phorbol ester (PMA) was added to a final concentration of 100ng / ml, and culture was continued for 48h to induce monocytes to differentiate into macrophages. The experimental group was added with 0.5 mg / ml and 1 mg / ml sulfated mannoglucuronide hexaose respectively. Culture for 48 hours, remove the medium, wash with PBS, add 100 μL of RIPA lysate containing protease and phosphatase inhibitors, scrape off the cells with a cell scraper, pipette the cell lysate into a 1.5mL EP tube and lyse the cells on ice for 30min; 4 Centrifuge at 13000rpm for 15min, and pipette the supernatant into a new 1.5mL EP tube. Concentration determination was performed with BCA protein concentration determination kit, and stored in a -80°C refrigerator. Western blot was...
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