| Mirabilis himalaica belongs to Mirabilis L.(Nyctaginaceae).Perennial herb,mainly distributed in high altitude areas of 3 300 m,such as Tibet,Qinghai,Gansu and other places,as well as northwest India.Mirabilis himalaica is lightloving and resistant to radiation,and is especially the ability to adapt to drought is strong.The dried root is used as medicine,which is part of the multi-flavored tibetan medicine and the traditional five roots.It has the function of warming kidneys and promoting muscle growth.Diuresis,stone-removal and other effects,treat stomach cold,kidney cold,lower body cold,tumor and other diseases.Polysaccharides are macromolecular substances connected by glycosidic bonds and have a wide range of biological activities.At present,as many as thousands of polysaccharide compounds have been isolated from natural products.At present,domestic and foreign scholars have isolated flavonoids,total saponins,trigonelline,total polyphenols,rotenone and polysaccharide compounds from Himalayan Mirabilis.In this experiment,the hot water extraction process in the Himalaica roots of Mirabiles is optimized by responding to the surface,the optimized process parameters are obtained.: the extraction time was 2 h,the extraction temperature was 85℃,and the solid-liquid ratio was 1:24 g/m L.At this time,extraction rate of polysaccharides was 13.1024%.The sugar content of refined polysaccharide MHPS-1 was 86.4%.The molecular weight and purity of MHPS-1 were analyzed by HPGPC,and it was found that the polysaccharide peak shape was single and symmetrical,and the polysaccharide is a homogeneous polysaccharide with a molecular weight of 28 495 Da..The composition of monosaccharose is analyzed by highperformance liquid chromatography to calculate the molar ratio..The result was fucose: rhamnose : arabinose : galactose : glucose : galacturonic acid : glucuronic acid = 0.013 : 0.097 : 0.086 : 0.07 : 0.006 : 0.701 : 0.028,indicating that MHPS-1 mainly contains galacturonic acid.Combining with infrared spectrum(FT-IR),methylation and nuclear magnetic resonance(NMR)to analyze the connection mode of monosaccharides and the type of glycosidic bonds,the results show that the main connection mode in MHPS-1 is → 4)-α – D – Galp A-(1→4)– α – D– Galp A-(1→4)– α – D – Galp A(6 – Ome)-(1→,It‘s a galacturonic acid polysaccharide.The MTT experiment was used to study the inhibitory effect of Mirabilis Himalaica polysaccharide MHPS-1 on tumor cell proliferation,and the apoptosis of cells was observed with a microscope.The results show that MHPS-1combined with cisplatin can inhibit the proliferation of human gastric cancer cells BGC-823,and the inhibition rate has a good dose-effect relationship with the concentration of polysaccharides and the duration of action.When 500 μg/m L MHPS-1 was used in combination with cisplatin,the inhibitory rate of BGC-823 cells was 51%,and the number of migrating cells and invasion were reduced by41.90% and 52.90%,separately.The Annexin V-FITC/PI double staining method and Tunel staining method detected that the apoptosis rate was increased by 19.28%and 16.30% compared with the blank group,respectively.The results of Western blot showed that after MHPS-1,the BGC-823 cell Bax the relative expression of Caspase-3(17KD)and Caspase-9(35 KD)protein increased,increased,the relative expression of Bcl-2 protein decreased.Fluorescence microscopy and flow cytometry detected that MHPS-1 and its cisplatin have increased the level of cellular reactive oxygen species and decreased mitochondrial membrane potential.The immunofluorescence detection of the expression of NF-k B p65 showed that MHPS – 1 combined with cisplatin was the average optical density value after incubation decreased by 46.8%. |
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