Purification, characterization and in vitro and in vivo immune enhancement of polysaccharides from mulberry leaves
Purification, characterization and in vitro and in vivo immune enhancement of polysaccharides from mulberry leaves
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桑叶多糖的纯化、表征及其体内外免疫增强作用
DOI：
10.1371/journal.pone.0208611
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发表时间：
2019-01-02
期刊：
影响因子：
3.7
通讯作者：
Jiang, Chunmao
中科院分区：
文献类型：
--
作者：
Chen, Xiaolan;Sheng, Zhicun;Jiang, Chunmao
桑叶多糖（MLP）通过DEAE-52纤维素和Sephadex G-100柱层析进行提取和纯化，得到两种主要的纯化多糖（MLP-1和MLP-2）。对纯化的多糖进行了表征，并研究了它们的免疫增强特性。 MLP-1的分子量为9.31×10(4)Da，由甘露糖、鼠李糖、葡萄糖、半乳糖、木糖和阿拉伯糖组成，摩尔比为0.71:1.00:2.76:1.13:3.70:2.81。 MLP-2的分子量为2.22x10(6)Da，其单糖成分为甘露糖、鼠李糖、葡萄糖、半乳糖和阿拉伯糖，摩尔比为1.31:8.45:6.94:1.00:11.96。红外光谱显示每个MLP具有典型的糖吸收峰特征，紫外(UV)光谱显示MLP均不含核酸或蛋白质成分。然后，通过3-[4,5-二甲基噻唑-2-基]-2,5-二苯基溴化四唑（MTT）测定比较这些多糖在体外刺激小鼠脾淋巴细胞增殖的能力。 MLP-2 比 MLP-1 更有效；因此，选择MLP-2来研究其体内免疫增强作用。体内实验采用新城疫（ND）疫苗免疫14日龄鸡口服MLP-2，并以黄芪多糖（APS）作为对照。每只鸡从ND疫苗免疫前三天开始连续7天口服给予4mg或8mg MLP-2。 MLP-2显着提高了ND血清抗体滴度以及气管和空肠冲洗液中白细胞介素2（IL-2）、干扰素γ（IFN-γ）和免疫球蛋白A（sIgA）浓度，并增加了盲肠扁桃体中免疫球蛋白A阳性（IgA（+））细胞的数量并增加了体重。这些结果表明MLP-2可以显着增强免疫活性，因此可以用作免疫增强剂候选药物。
Mulberry leaf polysaccharide (MLP) was extracted and purified by DEAE-52 cellulose and Sephadex G-100 column chromatography to afford two major purified polysaccharides (MLP-1 and MLP-2). The purified polysaccharides were characterized, and their immuneenhancing properties were investigated. MLP-1 had a molecular weight of 9.31x10(4) Da and was composed of mannose, rhamnose, glucose, galactose, xylose, and arabinose in a molar ratio of 0.71:1.00:2.76:1.13:3.70:2.81. The molecular weight of MLP-2 was 2.22x10(6) Da, and its monosaccharide constituents were mannose, rhamnose, glucose, galactose, and arabinose in a molar ratio of 1.31:8.45:6.94:1.00:11.96. Infrared spectroscopy showed that each MLP had a typical absorption peak characteristic of sugars, and ultraviolet (UV) spectroscopy showed that neither MLP contained nucleic acid or protein components. Then, the abilities of these polysaccharides to stimulate spleen lymphocyte proliferation in mice in vitro were compared by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. MLP-2 was more effective than MLP-1; therefore, MLP-2 was chosen for the study of its immune-enhancing effects in vivo. For the in vivo experiments, 14-day-old chickens immunized with Newcastle disease (ND) vaccine were orally administered MLP-2, and Astragalus polysaccharide (APS) was used as the control. Each chicken was orally administered 4 mg or 8 mg of MLP-2 for seven consecutive days starting three days before ND vaccine immunization. MLP-2 significantly improved the ND serum antibody titer and interleukin-2 (IL-2), interferon-gamma (IFN-gamma) and immunoglobulin A (sIgA) concentrations in tracheal and jejunal wash fluids, and increasing numbers of immune globulin A-positive (IgA(+)) cells in cecal tonsils and increased body weight. These results indicated that MLP-2 could significantly enhance immune activity and could therefore be utilized as an immunopotentiator drug candidate.