Structural Characterization Of A Polysaccharide From Arillus Longan And Its Effects On Tumor Cells And Immune Cells In Vitro

Objective: To extract the active polysaccharides from Dimocarpus Longan and purify Dimocarpus Longan Lour. polysaccharides and study their chemical structures and investigate the effects on tumor and immune systerm, in order to provide the theoretical basis for its reasonable development and use.Methods: Crude polysaccharide LYR was extracted respectively by methods of hot-water extraction and supersonic wave. The total sugar contents of crud polysaccharide LYR was determined by Phenol-sulfuric acid method. LYR was purified by deproteinizing of enzyme-Sevage method and protease enzymic hydrolysis method and decolouring by H2O2. LYR was further purified by DEAE-cellulose (OH-) ion-exchange and Sephacryl S-300 chromatographies.Monosaccharide moiety in LYRP2 was determined by HPGPC. The structrual characterizations of LYRP2 were investigated by the method of periodic acid oxidation and Smith degradation, part-acid hydrolysis and exhaustive methylation. The structural characterizations of LYRP2 were confirmed by gas chromatogram, infrared spectrum, gas-mass spectrum. The LYRP2 activities of anti-tumor and immune in vitro were assayed by MTT method.Results: The yield of hot-water extraction method was 3.4% and sugar content was 62%, but the method of supersonic wave were 7.0% and 64%respectively. After ten times of enzyme-Sevage method, rate of deproteination of LYR was 52%, and sugar expenses was 39%. But after twice of protease enzymic hydrolysis method, they were 84% and 20%, respectively. After decolouring, LYR were white or pallide-flavens. The homogeneous polysaccharide LYRP2 was first isolated from Arillus Longan, and the molecular weight of LYRP2 was 1.1×-105. The monosaccharide moiety of LYRP2 was analysized by complete acid hydrolysis and was D-glucose. Based on the analysis of IR spectrum, LYRP2 was pyranose with the glycoside configuration of P-hemiacetal hydroxyl. According to the study of structural characterizations,the results indicated that LYRP2 had a main chain ?-D- (1?3) -Glcp, and branches ?-D- (1?3) -Glcp-?-D-1) -Glcp, and ?-D- (1?4) -Glcp-?-D-1)-Glcp in which 1-linked Glcp were terminal residues, linked to the main chain.LYRP2-1 had inhibitory effect with more than 40% inhibitory ratio on SKV03 cell line at the concentration of 5?40 mg/L, IC50 was 5.6mg/L. LYRP2-2 had inhibitory effect with more than 30% inhibitory ratio on SKV03 cell line at the concentration of 5?40 mg/L, the highest inhibitory ratio was 39.9% at the concentration of 40mg/L. LYRP2-1 had inhibitory effect with more than 40%inhibitory ratio on H08910 cell line at the concentration of 320?40 mg/L, IC50 was 114mg/L. LYRP2-2 had inhibitory effect with more than 40% inhibitory ratio on H08910 cell line at the concentration of 320?40 mg/L, IC50 was 325mg/L. The experiments in vitro showed that LYRP2-1 and LYRP2-2 could proliferate spleen lymphocytes from normal BALB/c mice at the concentration of 3.2?400 mg/L, and the highest stimulation index were 124% and 140%respectively. LYRP2-1 could inhibit the proliferation response of spleen lymphocytes from normal BALB/c mice induced by ConA (5 mg/L) in vitro,and at the concentration of 3.2?400 mg/L the lowest stimulation index of LYRP2-1 was 68%. And LYRP2-2 could proliferate the proliferation response of spleen lymphocytes from normal BALB/c mice induced by Con A (5 mg/L) in vitro, and at the concentration of 3.2?400 mg/L the highest stimulation index of LYRP2-2 was 157%.Conclusion: Supersonic wave method is better than hot water extraction method. Protease enzymic hydrolysis method is better than enzyme-Sevage method. The homogeneous polysaccharide LYRP2 is first isolated from Arillus Longan and the molecular weight of LYRP2 is 1.1×105. The monosaccharide moiety of LYRP2 is D-glucose. LYRP2 is pyranose with the glycoside configuration of ?-hemiacetal hydroxyl. According to the analysis of chemical structure, the results indicate that LYRP2 has a main chain P-D- (1?3) -Glcp,and branches ?-D- (1?3) -Glcp-?-D-1) -Glcp,and ?-D- (1?4) -Glcp-?-D-1)-Glcp in which 1-linked Glcp are terminal residues, linked to the main chain.LYRP2-1 and LYRP2-2 have potent inhibitory effect on the growth of tumor cell lines of SKV03 and HO8910, and LYRP2-1 has showed cell toxic more intensively than LYRP2-2 at the same concentration. LYRP2-1 and LYRP2-2 can enhance the immune system functions and the biological activity of LYRP2-1 may not result from selecticely activating T cells.