| Abalone viscera and abalone muscle are rich in polysaccharides,which has good biological activity.In this study,abalone visceral polysaccharides（AVP）and abalone muscle polysaccharides（AMP）were extracted using protease hydrolysis combined with ethanol fractional precipitation.The physicochemical properties of two crude polysaccharides and their purified components were studied.And the primary structure of abalone muscle polysaccharides was explored.Finally,the effects of AVP and AMP on mouse peritoneal macrophages were investigated.Firstly,AVP and AMP were separated by DEAE-52 anion exchange column chromatography,and their physicochemical properties were investigated and compared.The contents of carbohydrate,protein,uronic acid and sulfate in AVP were 58.6%,8.3%,15.0%and13.3%,respectively.Three fractions were obtained by DEAE-52 column chromatography from AVP and AMP,respectively.Based on the results of chemical composition,monosaccharide and FTIR analysis,it was found that AVP was mainly composed of two acidic polysaccharides（AVP-2 and AVP-3）.Compared to the AVP,although the carbohydrate content in both AVP-2 and AVP-3 did not increased,the main monosaccharide composition of Gal was increased from47.4%to 69.2%.On the other hand,the contents of carbohydrate,protein,uronic acid and sulfate in AMP were 83.9%,3.6%,3.5%and 4.5%,respectively.Three fractions recovery rates of AMP-1,AMP-2 and AMP-3 from AMP were 63.0%,8.6%and 6.1%,respectively.Based on the results of monosaccharide analysis,it was found that AMP was mainly composed of Glc in molar ratio of 92.6%.After separated from AMP,the main monosaccharide composition of Glc was increased in AVP-1,but decreased in both AMP-2 and AMP-3.Based on above results and FTIR analysis,it was found that both AVP-1 and AMP-1 belonged to neutral polysaccharides,while other fractions belonged to acidic polysaccharides.Furthermore,both two neutral polysaccharides of AVP-1 and AMP-1 showed good antitumor activity in vitro.Nextly,in order to understand the biological activity of polysaccharides and their relationship,AMP-1 was selected for further structural studies due to its good antitumor activity and simple monosaccharide composition.AMP-1 was purified by Sephadex G-25 column chromatography,and a homogeneous polysaccharide（AMPP）was obtained with the molecular weight of 3.2 kDa.AMPP was composed of Glc and Man in a molar ratio of 99.7:0.3.The results of methylation showed that AMPP was mainly composed of→4)-α-D-Glcp-（1→,→4,6）-α-D-Glcp-（1→,→6）-α-D-Glcp-(1→andα-D-Glcp-(1→in molar ratio of 52.44:17.74:7.24:22.58.Based on the results of FTIR analysis,methylation analysis and 1D(1H NMR and13C NMR)and 2D（COSY,HSQC,HMBC and ROESY）NMR experiments,the obtained AMPP was determined to be anα-1,4-,1,6-linked D-glucan.In one repeat unit of AMPP structure,the backbone chain was contained of eight 1→4 glycosidic bond and one 1→6 glycosidic bond,with three branch chains linked by 1→6 glycosidic bond.Finally,the effects of AVP and AMP on mouse peritoneal macrophages were investigated.The results show that both AVP and AMP could inhibit the phagocytic activity of mouse peritoneal macrophages and LPS-induced peritoneal macrophages,and have dose-related.AVP could also inhibit the release of NO from macrophages and LPS-induced macrophages by down-regulating the activity of iNOs,and has dose-dependent,but AMP had no significant effect on iNOs activity and the release of NO.In addition,AVP and AMP could effect the TNF-αsecretion levels of the mouse peritoneal macrophages.With the increase of AVP concentration,the secretion of TNF-αin mouse peritoneal macrophages and LPS-induced mouse peritoneal macrophages showed a tendency to decrease first and then increase.The optimal inhibitory concentrations were 500μg/mL and 250μg/m L,respectively.With the increase of AMP concentration,the levels of TNF-αin mouse peritoneal macrophages and LPS-induced murineperitoneal macrophages were gradually increased.Compared with AMP,AVP has a better anti-inflammatory effect. |
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