Isolation,Purification,Structural Identification And Anti-alzheimer’s Disease Activity Of Saccharides From Alpinia Oxyphylla

This article takes saccharides of Alpinia oxyphylla as the research object,and its anti-Alzheimer’s disease activity in vivo has been thoroughly investigated.In addition,the crude saccharides of A.oxyphylla were further separated and purified to obtain homogeneous saccharides,and the structures were elucidated.Four kinds of crude sugar polymers AO50,AO70,AO90 and AOB were obtained by water extraction and alkali extraction and alcohol precipitation.Mice model of Alzheimer’s disease(AD)were successfully constructed by intraperitoneal injection of 1 mg/kg scopolamine hydrobromide.Behavioral experiments were used to evaluate the effect of A.oxyphylla crude saccharides on learning and memory ability and anxiety behaviors in AD model mice.The results showed that the A.oxyphylla crude saccharides(500 mg/kg)significantly improved the learning and memory ability and anxiety behaviors of AD mice,which compared with the positive drug Huperzine A(2 mg/kg),there was no significant difference.In the detection of biochemical indicators,the A.oxyphylla crude saccharides markedly reduced the levels of NO,IL-6,IL-1β,TNF-α and PGE-2 in brain tissue and serum of AD mice,increased the levels of GSH and SOD in mice,and reduced the MDA levels.It is indicated that the A.oxyphylla crude saccharides may play an anti-Alzheimer’s disease role by regulating the expression of inflammatory factors and oxidative stress factors.Metabolomics studies using gas-mass spectrometry showed that the crude saccharides AO50,AO70,AO90 and AOB can regulate the content of biomarkers in AD mice,which adjust their metabolite levels closer to the normal group,which further showed that the A.oxyphylla crude saccharides have significant anti-Alzheimer’s disease activity.Based on the positive anti-Alzheimer’s disease activity in vivo,the A.oxyphylla crude saccharides were deproteined,dialyzed,lyophilized and isolated via DEAE-Cellulose 52 and Sephadex G-75 column chromatography.Seven homogeneous saccharides AOP50-2-1,AOP70-1,AOP70-2-1,AOP70-2-2,AOP70-3-1,AOP70-3-2 and AOP90-2 were obtained.Specific optical rotation method and HPGPC showed that they were homogeneous saccharide with relative molecular weights of 72.45 k Da,5.33 kDa,76.66 kDa,16.53 kDa,36.85 kDa,12.20 kDa,and 1.83 kDa,respectively.Pre-column derivatization-HPLC was used to determine the monosaccharide composition.Methylated-GC-MS was used to determine the types and proportions of sugar residues.The 1D and 2D NMR techniques were used to analyze the connection of each sugar residue site.The structures of seven homogeneous saccharides were identified,respectively.The results showed that AOP50-2-1,AOP70-1,AOP70-2-1,AOP70-2-2,AOP70-3-1,AOP70-3-2 and AOP90-2 were all acid heteropolysaccharides.The surface of AOP70-1 had irregular circular depressions and rough texture;AOP70-2-1 had an irregular sheet structure with rough wrinkles on the surface;AOP70-2-2 is an irregular sheet-like structure with a smooth surface;AOP70-3-1 is a loose three-dimensional structure formed by stacking multiple sheets;AOP70-3-2 is a filament-like structure,distributed scattered,rough surface with wrinkles;AOP90-2 has a chip-like structure with small holes on its surface.The structures of the seven homologous saccharides have not been reported in previous literature.