| Diabetes Mellitus is a kind of endocrine metabolic disease. Due to the lack of effective medications during the long progression period, it has become one of the major diseases that threaten human health. The research on the anti-diabetic activity of polysaccharides of Anoectochilus roxburghii (PSA) could help to find a safe and reliable drug for the prevention of diabetes. At present, the research on finding the active anti-diabetic ingredients of Anoectochilus roxburghii and the underlying mechanism is still in the initial stage. In addition, the molecular structure of the active ingredients is rarely reported.In this study, polysaccharides from Anoectochilus roxburghii (PSA) were extracted, and their structures and anti-diabetic effects were investigated. In vitro experiments were carried out to detect the antioxidant capacity and the inhibition ofα-glucosidase by PSA. In vivo experiments were performed to determine the anti-diabetic activity of PSA. The possible underlying mechanisms were discussed in the end.The experiment includes four parts:(1) Extraction and purification of PSA Raw PSA was extracted using the ultrasound-assisted extraction technology. PSA-1 and PSA-2 were further purified from PSA using ion-exchange and glucose gel column chromatography. The redults showed that PSA-1 was a proteoglycan with a molecular weight of 14208 Da. The protein content of PSA-1 was 0.44%. On the other hand, PSA-2 was a polysaccharide with a molecular weight of 17787 Da. No protein was found in PSA-2.(2) Structual analysis of PSA The results from infrared spectrum (IR) analysis showed that the absorption peaks were concordant for the functional groups, but disparated in the fingerprint region of PSA-1 and PSA-2. Gas chromatography (GC) analysis demonstrated that PSA-1 was composed of rhamnose, arabinose, xylose, mannose, galactose and glucose; PSA-2 was composed of galactose and glucose. The structural analysis using chemical methods and UV-spectrometry, such as periodic acid oxidation and I2-KI reaction, indicated that the monosaccharides were connected by 1â†'2,1â†'2,6,1â†'4,or 1â†'4,6 bonds. No triple helix structure was found in PSA-1 and PSA-2. The results also revealed the existence of long side chains and aboundent branch chains. The protein of PSA-1 was connected by O-link polysaccharide.(3) In vitro experiment The results showed that PSA, PSA-1 and PSA-2 had a strong removing and reducing ability for radicals including DPPH·,·OH, O2-·. PSA possessed an inhibitory activity againstα-glucosidase with an IC50 of 27.01μg/mL.(4) In vivo experiment Alloxan-induced diabetic mice were treated intragastrically with different doses of PSA. The administration of PSA for 14 days caused an apparent decrease in blood glucose levels (p<0 .05); Simultaneously, the concentrations of serum total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) were decreased, while the levels of high-density lipoprotein cholesterol (HDL-C) were apparently increased. In addition, PSA caused a significant decrease (p<0.05) in the level of malondialdehyde (MDA) and increase (p<0.05) in the enzymic activities of antioxidants (SOD, GSH-Px and T-AOC) in sera and tissues of diabetic mice. These effects were dose-related to PSA. The tissues (pancreas, liver, kindey and heart) were stained with hematoxylin/eosin. The results from histopathology showed that tissue damage was decreased or repaired.In conclusion, the results from this study demonstrated that the anti-diabetic effect of PSA was resulted from its anti-oxidative, anti-dislipidemic and tissue-repairing capabilities. Therefore, Anoectochilus roxburghii could have a good application value in the prevention of diabetes. |
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