Isolation, Purification And Hepatoprotective Effect Of Polysaccharides From Dicliptera Chinensis

The dried aerial part of D. chinensis is one kind of both food and oriental folkmedicine that has been traditionally used for years to prevent hyperpyrexia anddetoxication. This paper focuses on the isolation, purification, structurecharacterization and hepatoprotective effect of the polysaccharides in Diclipterachinensis. The chief results are showed as follows:1. Explore the effective part of Dicliptera chinensis polysaccharide(DCP).Totalpolysaccharides was extracted from Dicliptera chinensis. Mw<2kD、Mw<6kD、Mw6Kd-10kD、 Mw>10kD group was divided by dialysising. Establishedparacetamol(APAP), aclohol and D-galactosamine(D-GalN) induced acute liverinjury model. Serum and liver indicators were determined. Combined with hepatichistopathological examination to determine the effective parts. The results follows:(1) In the APAP model: campared with model group, DCP prior to theadministration of APAP significantly prevented the increases of alanineaminotransferase (ALT) and aspartate transaminase (AST) level in serum(P<0.01orp<0.05), and malondialdehyde (MDA) accumulation, glutathione (GSH) anduperoxide dismutase (SOD) depletion in liver tissue(P<0.01or p<0.05).(2) In the alcohol model: campared with model group, DCP prior to theadministration of APAP significantly prevented the increases of ALT and ASTactivities in serum(P<0.01or p<0.05), and malondialdehyde (MDA) accumulation,improved glutathione (GST) depletion in liver tissue(P<0.01or p<0.05).(3)In the D-GalN model: campared with model group, DCP prior to theadministration of APAP significantly prevented the increases of ALT and ASTactivities in serum(P<0.01or p<0.05), and reduced malondialdehyde (MDA)accumulation, increase glutathione (GST)activity in liver tissue(P<0.01or p<0.05).Our results indicated that DCP can protect against APAP and alcohol-inducedacute liver injury in mice, and the most active part is the range of molecular weightleft than6kD.2. Study in liver fibrosis of the molecular weight less than6kDa parts ofDCP.Intraperitoneal injection of carbon tetrachloride in rats, once every three days,lasting8weeks to establish liver fibrosis model. The establishment of the normal group, model group, the glycyrrhizin group (30mg/kg), Fufangbiejiarangan liverslices (600mg/kg), the molecular weight less than6kDa group (120mg/kg),glycyrrhizin and polysaccharide combination group (150mg/kg), administered dailyfor8weeks, the rats were sacrificed, specimens from rat liver and serumdetermination of relevant indicators. The results showed: Compared with model group,the treatment group could significantly reduce serum ALT, AST, HA, LN,PCⅢcontent (p <0.01) and reduced liver tissue homogenates in Hyp, TGF-β1content(p <0.01or p <0.05). Histopathological examination by HE staining and Massonstaining, the results of drug-treated liver fibrosis had different degrees ofimprovement, fiber spacing was narrowed, connective tissue proliferation reducedinflammatory infiltration lessened. The combination group had the best curative effect,steatosis disappeared, tending to be normal liver tissue structure.3. Purification of DCP. DCP by the DEAE-52anion exchange column, gradientelution, separation of the four components: P1, P2, P3, P4. The four components werefurther purified by Sephadex G-50gel column chromatography, respectively. Thenfour sub-components were receiving: P1A, P2A, P3A, P4A. t of no four components.The UV spectrum of260nm,280nm has no absorption described that each componentexcluding protein.Molecular weight of each sub-components was determinate by gelpermeation chromatography (GPC). The molecular weight are respectively::2339Da.The monosaccharide composition results showed that: P1A and P3A contain the6kinds of monosaccharides: rha, ara, xyl, man, glu, gal; P2A comprising5kinds ofmonosaccharides: rha, ara, man, Glu, gal; the P4A containing4kinds ofmonosaccharides: rha, man, glu, gal.4. The P1A component structural analysis. Partial acid hydrolysis, periodate oxidationand Smith degradation, IR,1H-NMR and13C-NMR analysis showed that: The mainchain of monosaccharide residues of P1A were rhamnose, mannose, glucose andgalactose, the terminal residue and branch monosaccharide residue as arabinose andxylose. P1A containing1-,1,6-bond type and the non-reducing end which couldproduce formic acid, and also containing l,2-and1,4-bond type which couldconsume NaIO4and not produce formic acid, two key-type ratio is about1:2. Theoxidizable key type accounted for45%of non-oxidizable key type mainly1,3-keysaccounted for55%of.