Preparation Of Polysaccharide From The Brown Seaweed Sargassum Graminifolium,and Its Inhibition Effects On Calcium Oxalate Stones

Urinary stones affect a large proportion of the population, most of urinary stones are calcium oxalate stones. Therefore, it is important to search for new bioactive compounds to meet the clinical needs that prevent calcium oxalate stones and reduce their recurrence rate. Sargassum graminifolium, a brown seaweed extensively distributed along the coasts of the South and the East China Sea. This thesis researched on preparation of polysaccharides from Sargassum graminifolium (SGP), and its physical and chemical properties, antioxidant effects and also the inhibition of calcium oxalate crystals activity in vitro, furthermore the treatment efficacy and mechanism of SGP on renal injury induced by hyperoxaluria in vivo. The main view is based on antioxidant effects, reduction of calcium oxalate crystal deposition and protection mitochondrial function. The summary as follows:Firstly for SGP preparation technology, considering pharmaceutical and health care applications, the processes of ultrasonic assisted extraction, deproteinization and decolorization were used, with the methods of single factor and orthogonal array design. In order to identify SGP’s physical and chemical properties, the methods include electrophoresis, ultraviolet spectroscopy, infrared spectroscopy, mass spectrometry and thin-layer chromatography were used, got the content of sulfate, glucuronic acid and protein (sulfate13.2%, glucuronic acid10.11%, protein1.03%), and also molecular weight of SGP was obtained(11887Da). We can conclude that SGP is a kind of sulfated polysaccharide composed of glucose, galactose and mannuronic acid.Secondly, for analyzing the antioxidant activity of SGP in vitro, the reducing power, the ability of scavenge superoxide radicals, and the activity of scavenge DPPH were determinted respectively. The results showed there is obvious dose-effect relationship between concentrations of SGP and the three antioxidant activities, increasing concentrations of SGP resulted in increased scavenging of superoxide anions and DPPH radicals (IC50=1.9and0.56mg/mL, respectively).Thirdly this thesis researched on the effects of SGP on calcium oxalate crystallization in vivo. We studied the maximum time, the rates of nucleation and aggregation that characterize the crystallization process and also investigated the morphology of calcium oxalate crystals. Furthermore, the Zeta potential measurement of calcium oxalate crystals liquids was tested after adding different concentrations of SGP in order to find the relationship between concentrations of SGP and the negative charge of calcium oxalate particle surface. The results show that SGP’rates of nucleation and aggregation were69.2%and76.8%, and it’s better than with trisodium citrate.At last, this thesis researched the effects of SGP on hyperoxaluria rats and its action mechanisms. For the rats with calcium oxalate stones, they were taken high, middle and low dose of SGP. On following aspects the rats were measured:the urine physiological indicators, the biochemical serum markers, the kidney mitochondria indicators (the activity of ATP enzyme, GSH-PX enzyme, SDH enzyme and SOD enzyme and mitochondrial swelling, MDA levels), and also renal morphology was investigated using tissue slice method. Base on these experiments, this thesis researched SGP protects the kidney cells of hyperoxaluria rats from damaging through antioxidant, reducing calcium oxalate crystal deposition, and protection mitochondria. Then it discovered the vital mechanism of SGP on the inhabition activity of calcium oxalate stones, for it has obvious antioxidant ability, and can regulate the mitochondrial dysfunction of rat kidney.