| Sargassum fusiforme,Sargassum family,alias sea vegetable bud,staghorn,etc.,had a long history of edible and medicinal in China.There have been reports on hypoglycemic studies on Sargassum fusiforme,but most of them verify the efficacy of the extracted ingredients by using animal experiments.It is rare to screen active ingredients from Sargassum fusiforme from the perspective of sugar enzyme inhibition,especially to effectively inhibit?-glucosidase.In this study,the polysaccharide component that can effectively inhibit?-glucosidase was isolated and purified from Sargassum fusiforme,to explore its inhibitory effect on?-glucosidase and to conduct preliminary analysis on the physical and chemical properties and structural characteristics of the active component.Provide new ideas for the development of food-borne hypoglycemic components.The main research results are as follows:1.Using Sargassum fusiforme as raw material,the alkaline protease assisted water extraction and alcohol precipitation method was used to extract Sargassum fusiforme polysaccharides.From the single-factor experiment results,The optimal parameters for extraction of hijiki polysaccharides were:extraction temperature 50?,enzyme addition 0.9%,pH 10,substrate concentration 4%,extraction time 4 h.Under thses conditions,the extraction rate of crude polysaccharides from Sargassum fusiforme was 11.51%.The acid polysaccharide SFP-1 was isolated and purified from Sargassum fusiforme using DEAE-52 anion exchange column and Sephadex G-200 gel column chromatography.2.The results of inhibition experiments on?-glucosidase activity showed that the half-inhibitory concentration of SFP-1 was 0.681 mg/mL,which was better than acarbose(1.308mg/mL).At the same time,SFP-1 had good temperature and acid-base stability,and can maintain high inhibitory activity at 30?100?and p H 3?10.And it can maintain good inhibitory activity in simulated gastrointestinal fluid.Preliminary kinetic studies revealed that it was a reversible mixed inhibition.The interaction between SFP-1 and?-glucosidase was analyzed by fluorescence spectroscopy,CD chromatography and ultraviolet spectroscopy.It was found that SFP-1 exhibited a static quenching effect on the intrinsic fluorescence of?-glucosidase.The binding process induced the conformational change of?-glucosidase.These results indicated that SFP-1 may become a novel?-glucosidase inhibitor for functional foods.3.Chemical structure analysis showed that the average molecular weight of SFP-1 was160.63×10~3,mainly composed of fucose,galactose,glucose,xylose,mannose and glucuronic acid,and its relative molar ratio is 15.17:7.72:4.92:1.99:4.04:23.07.The sulfate group content of SFP-1 was 3.04%.The results of Fourier transform infrared spectroscopy and nuclear magnetic analysis showed that SFP-1 had both?-type and?-type glycosidic bonds in the sugar chain.From the results of periodate oxidation,it can be seen that the SFP-1 sugar chain may contain 1?or 1?6 glycosidic bonds,1?2 or 1?4 glycosidic bonds;from Smith degradation products,it can be known that the SFP-1 sugar chain must contain 1?4 Glycosidic bond.4.Thermogravimetric analysis shows that SFP-1 had good thermal stability.Atomic force microscopy showed that SFP-1 in solution existed in an irregular particle structure,and the height of the single polysaccharide chain is about 0.5-1.5 nm higher than that of a single polysaccharide chain(0.1-1 nm).These results indicated that SFP-1 may have branching and entanglement,and there was aggregation in polysaccharide molecules.X-ray diffraction and Congo red experiments showed that SFP-1 had an amorphous structure and no three-helix spatial conformation. |
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