| Gelidium amansii belongs to the phylum of red algae,subclass of true red algae,order of Stonehullidae,family of Stonehulaceae,genus of Caragana.Most of the Gelidium amansii is red and widely spread over coast area.Gelidium amansii is one of the mainly raw material for making agarose,and it is widely used in medicine and food.It has the function of clearing heat,detoxifying,anti-coagulation,lowering blood pressure and reducing lipid.The substance extracted from Gelidium amansii by alcohol has anti-bacterial and anti-oxidant activity.In recent years,Algal polysaccharides have attracted more attention of researchers all over the world due to its biological activities.However,there are few reports on Gelidium amansii polysaccharides(GAP).In this paper,extraction,separation and purification,physicochemical and structural characterization and biological activity of GAP were studied and analyzed.The contents in detail were as follows:1、Study on extraction parameters of GAPExtracting time,temperature,the ratio of material to liquid and times are major factors that affected the rate of extracting GAP.These factors were analyzed by single factor experiments.And three of them(extracting temperature,extracting time and liquid-material ratio)were selected to be analyzed via the Box-Behnken design Three factors and three horizontal response surface optimization experiments were adopted,and the data were statistically analyzed by Design-Experts 8.0.6 software.The optimal extracting conditions of GAP were obtained as follows:extracting temperature 62.11℃ extracting time 1.6 h,Material to liquid ratio 35.55 mL/g.According to the conditions,the yield of GAP is 4.17%.2、Isolation and purification,Physicochemical and Structural Characterization of GAPIn this experiment,crade Gelidium amansii polysaccharides(Crude GAP)was purified by DEAE-Cellulose Anion Exchange Chromatography column,and four main compoents(GAP-1,GAP-2,GAP-39 GAP-4)were obtained.Determination of Crude and purified GAP were perfomed according previous methods with slightly modification.The results showed that the carbohydrate contents of Crude GAP,GAP-1,GAP-2,GAP-3 and GAP-4 determined by phenol sulfuric acid method were 62.64±0.03%,67.16±0.01%,64.90 ± 0.04%,58.32 ±0.02%,47.30±0.04%,respectively.Protein contents of them were 3.36±0.02%,1.54±0.01%,0.99±0.01%,1.73±0.02%,2.02 ± 0.01%,respectively.Uronic acid contents determined by m-hydroxybiphenyl method were 4.29 ± 0.41%,2.88±0.30%,4.67±1.0%,4.98±0.38%,6.87 ± 0.73%,respectively.Sulfuric radical contents determined by barium chloride-gelatin method were 2.29±0.04%,1.29±0.11%,2.02±0.05%,3.46±0.37%,3.65±0.12%,respectively.Polyphenol content determined by Folin-Ciocalteu assay were too little.The monosaccharide composition of GAP were deternined by high performance liquid chromatography(HPLC),and the results were as follows:molar ratios of Crude GAP was Rib:Rha:Glc:Gal:Fuc=0.12:0.78:0.24:22.58:4.56,for GAP-1 were Gal:Fuc＝8.72:10.62,for GAP-2 were only Gal,and GAP-3 were GlcA:Gal=0.17:13.24,as well as GAP-4 were GlcA:Gal=3.02:23.61.The molecular weight of GAP-1,GAP-2,GAP-3 and GAP-4 were determined by HPLC.Results showed that the molecular weight of the four components were 1593 kDa,789 kDa,1225 kDa,1217 kDa,respectively.3、Simulated digestion in vitro of Crude GAPThe digestion simulated saliva,gastric juice and gastrointestinal juice under the in vitro was carried out for studing the digestion situation of Crude GAP.The results showed that Crude GAP had no significant change in molecular weight and reducing sugar content after digestion under saliva,simulated gastric juice and gastrointestinal juice.Besides,no free monosaccharides was detected in this process.The results indicated that Crude GAP was not degraded in oral,gastric juice and gastrointestinal juice.4、Antioxidant and immunomodulatory activities of GAPBy determining the index of eliminating DPPH radical,superoxide anion radical,ABTS radical,metal ion chela-ting force,reducing power and anti-lipid peroxidation,the antioxidant activity of Crude GAP and its two main purified components(GAP-2 and GAP-3)were studied.Both C-GAP,GAP-2,and GAP-3 had weak ability of scavenging DPPH radical,ABTS radical,However showed the strong scavenging activity of superoxide anion and significantly inhibited the lipid oxidation.The in vitro immunological activity of Crude GAPand its separated components GAP-2、GAP-3 were analyzed by cell model.The results show that Grade GAP and GAP-2、GAP-3 could effectively promote the proliferation of murine peritoneal macrophage RAW264.7 cell,and also improve the ability of macrophage phagocytosis and acid phosphatase,at the same time,it could effectively promote the release of NO from RAW264.7. |
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