巴戟天多糖分离、理化性质及对人肝细胞影响

Separation and physicochemical properties of polysaccharides from Morinda officinalis and their effects on human liver cells

目的对巴戟天粗多糖MOP-60和MOP-60-100进行分离和纯化获得多糖组分和成分,测定其理化性质,研究其对人LO2细胞的增殖和抗损伤作用。方法 MOP-60和MOP-60-100分别采用DEAE-纤维素柱色谱分离得到MOP-60-A、MOP-60-B、MOP-60-100-A、MOP-60-100-B和MOP-60-100-C共5个多糖组分;MOP-60-A又经透析获得均一多糖MOP-60-I;MOP-60-100-B又经Sephadex G-100柱色谱分离和纯化,得到MOP-60-100-Ⅰ和MOP-60-100-Ⅱ2个均一多糖。采用凝胶渗透色谱法测定多糖组分的相对分子质量分布和成分的峰高相对分子质量;采用酸水解、PMP衍生及毛细管电泳法测定单糖组成;采用MTT法,评价这些多糖组分和成分对人LO2肝细胞的细胞增殖及抗ConA(concanavalin A)和H2O2诱导LO2细胞损伤的作用。结果MOP-60-A、MOP-60-100-A和MOP-60-Ⅰ均由果糖(果聚糖)组成;MOP-60-Ⅰ峰高相对分子质量为2339;MOP-60-B、MOP-60-100-B、MOP-60-100-C 3种组分和MOP-60-100-Ⅰ和MOP-60-100-Ⅱ2种成分均是由多种单糖组成的杂多糖;MOP-60-100-Ⅰ和MOP-60-100-Ⅱ峰高相对分子质量分别为62 828和7783。体外活性评价结果显示,与溶剂组相比,MOP-60-B、MOP-60-100-B和MOP-60-100-C在250 mg/L浓度下显示明显的促肝细胞增殖活性(P<0.05);与ConA组相比,显示对肝细胞有明显的保护作用(P<0.01);仅MOP-60-100-C在100和250 mg/L浓度对H2O2引起的损伤显示较强的保护作用(P<0.05和P<0.01);多糖成分MOP-60-Ⅰ有一定促肝细胞增殖活性(P<0.05),而MOP-60-100-Ⅰ和MOP-60-100-Ⅱ作用不明显。结论巴戟天中酸性较强的多糖组分MOP-60-100-B和MOP-60-100-C对肝细胞有明显的促增殖和抗损伤作用。

Objective To separate the fractions and components of crud polysaccharides MOP-60 and MOP-60-100 from Morinda officinalis,characterize their physicochemical properties and effects on the LO2 cell proliferation as well as on the ConA-and H2O2-induced LO2 cell death.MethodsThe five fractions(MOP-60-A,MOP-60-B,MOP-60-100-A,MOP-60-100-B and MOP-60-100-C)were obtained by column chromatographic separation of MOP-60 or MOP-60-100 on a DEAE-DEAE-cellulose column.A component MOP-60-Ⅰwas obtained by dialysis of MOP-60-A.The further separation and purification of MOP-60-100-B by the Sephadex G-100 column chromatography afforded the other two components MOP-60-100-Ⅰand MOP-60-100-Ⅱ.The molecular distribution was determined with gel filtration chromatography.The monosaccharide compositions were analyzed with capillary electrophoresis after acid hydrolysis and PMP derivation.The effects of the fractions and components of polysaccharides on the human LO2 liver cell proliferation and on the ConA-and H2O2-induced LO2 cell damage were evaluated by the MTT method.ResultsMOP-60-A,MOP-60-100-A and MOP-60-Ⅰwere composed of fructose(fructosan),and the peak relative molecular mass of MOP-60-Ⅰwas 2339.MOP-60-B,MOP-60-100-B,MOP-60-100-C,MOP-60-100-Ⅰand MOP-60-100-Ⅱwere composed of multiple monosaccharides and heteroglycans.The peak relative molecular mass of MOP-60-100-Ⅰand MOP-60-100-Ⅱwere 62828 and 7783,respectively.MOP-60-B,MOP-60-100-B and MOP-60-100-C increased human LO2 hepatocyte proliferation and reduced the ConA-induced LO2 cell death at250 mg/L(P<0.01,compared to solvent or ConA alone group).MOP-60-100-C also reduced the H2O2-induced LO2 cell death at 100 and 250 mg/L(P<0.05 and P<0.01,compared to H2O2 alone group).ConclusionThe acidic fractions MOP-60-100-B and MOP-60-100-C from M.officinalis significantly promoted LO2 cell proliferation and inhibited the ConA-and H2O2-induced cell damage in human liver LO2 cells.