Page 104 - 2019年2月第30卷第4期

返魂草多糖中免疫增强活性有效组分的筛选及表征 Δ

周婷婷 ，朱迪夫 ，纪圣君 ，王春驰 ，贾东旭 ，李艳茹 ，唐 燕 （1.吉林省吉测检测技术有限公司，长春
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130117；2.吉林益民堂制药有限公司，吉林 四平 136001）
中图分类号 R932；R392.5 文献标志码 A 文章编号 1001-0408（2019）04-0523-05
DOI 10.6039/j.issn.1001-0408.2019.04.18
摘 要 目的：筛选并表征返魂草多糖组分中具有免疫增强活性的有效组分。方法：通过水提醇沉法提取返魂草浸膏（SCHE）多
糖并制得50％醇沉样品（SCHE-1）和80％醇沉样品（SCHE-2）。将小鼠单核巨噬细胞系RAW264.7细胞分为空白组（不含血清培
养基）、阴性对照组（含血清培养基）、脂多糖组（LPS，阳性对照，1 μg/mL）以及SCHE-1、SCHE-2低、高剂量（0.5、1 mg/mL）组。采
用四甲基偶氮唑盐（MTT）法检测RAW264.7细胞的细胞活性，采用酶联免疫吸附测定法（ELISA）检测RAW264.7细胞中白介素1β
（IL-1β）、IL-6、肿瘤坏死因子α（TNF-α）水平，以考察SCHE-1和SCHE-2对RAW264.7细胞的免疫增强活性。采用分子排阻色谱法
测定SCHE-1的分子量及其分布；采用高效液相色谱（HPLC）柱前衍生化法测定SCHE-1的单糖组成。采用NaOH法对SCHE-1进
行甲基化分析。结果：与阴性对照组比较，LPS 组及 SCHE-1 各剂量组均可显著增强 RAW264.7 细胞的活性并显著提高细胞中
IL-1β、IL-6、TNF-α的水平（P＜0.01），SCHE-1是具有免疫增强活性的有效组分。SCHE-1含中性糖40.05％、糖醛酸35.62％、蛋白
质8.89％；SCHE-1为混合物，分子量范围为62～6 119 Da，单糖组成主要为半乳糖醛酸、阿拉伯糖和半乳糖；SCHE-1的中性糖连接方
式以半乳糖的1→3、1→4和1→6连接为主，在1→3连接的O-6位上有分支，非还原末端主要为阿拉伯糖。结论：SCHE-1可能是返魂
草免疫增强活性的有效组分，其主要由多糖类成分组成；SCHE-1可能通过激活巨噬细胞释放IL-1β、IL-6、TNF-α而发挥调节机体免疫
功能的作用。
关键词 返魂草；多糖；免疫增强活性；筛选；表征

Screening and Characterization of Effective Components of Immunopotentiating Activity in Senecionis
cannabifolii Herba
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ZHOU Tingting ，ZHU Difu ，JI Shengjun ，WANG Chunchi ，JIA Dongxu ，LI Yanru ，TANG Yan（1. Jilin Jice
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Detective Technical Co.，Ltd.，Chuangchun 130117，China；2. Jilin Yimintang Pharmaceutical Co.，Ltd.，Jilin
Siping 136001，China）
ABSTRACT OBJECTIVE： To screen and characterize effective components of immunopotentiating activity in Senecionis
cannabifolii Herba. METHODS：The polysaccharide components were obtained by water extraction and alcohol precipitation
method to yield 50％ alcohol precipitation sample（SCHE-1）and 80％ alcohol precipitation sample（SCHE-2）. The cells from
mice mononuclear macrophage line RAW264.7 were divided into blank group（medium without serum），negative control group
（medium with serum），lipopolysaccharide group（LPS，positive control drug，1 μg/mL），SCHE-1 and SCHE-2 low-dose and
high-dose groups（0.5，1 mg/mL）. The cell viability of RAW264.7 cells was detected by MTT assay. The levels of IL-1β，IL-6 and
TNF-α in RAW264.7 were detected by ELISA. These were used to investigate the effects of SCHE-1 and SCHE-2 on the
immunological enhancing activity of RAW264.7 cells. The molecular weight and distribution of SCHE-1 were determined by size
exclusion chromatography；the monosaccharide composition of SCHE-1 was determined by HPLC pre-column derivatization.
Methylation analysis of SCHE-1 was conducted by NaOH method. RESULTS：Compared with negative control group，the activity
of RAW264.7 cells was enhanced significantly in SCHE-1 groups and LPS group，which also significantly increased the levels of
IL-1β，IL-6 and TNF-α in cell culture fluids（P＜0.01）. SCHE-1 was an effective component with immunopotentiating activity. The
neutral sugar content of SCHE-1 was 40.05％，the uronic acid was 35.62％，and the protein was 8.89％. SCHE-1 was a mixture，
molecular weight of which was 62-6 119 Da；monosaccharide was mainly composed of galacturonic acid，arabinose（Ara）and
galactose（Gal）. The results of methylation analysis showed that the backbone was composed of 1→3，1→4 and 1→6 linked Gal，
and branches were on the O-6 position of the 1→3 linked Gal，and the non-reducing terminals were Ara. CONCLUSIONS：
Δ 基金项目：吉林省科技发展计划项目（No.20180201020YY、 SCHE-1 may be the effective component of immuno
20180623037TC） potentiating activity， and main component of SCHE-1 is
＊主管药师，硕士。研究方向：中药新剂型与新技术。电话： polysaccharide. SCHE-1 may regulate the immune function by
0431-86172337。E-mail：zhoutingt2008@163.com activating macrophages to release IL-1β，IL-6 and TNF-α.
# 通信作者：主管药师。研究方向：中药活性成分与质量控制。 KEYWORDS Senecionis cannabifolii Herba；Polysaccharides；
电话：0431-86172337。E-mail：tangyan790601@126.com Immunopotentiating activity；Screening；Characterization

中国药房 2019年第30卷第4期 China Pharmacy 2019 Vol. 30 No. 4 ·523 ·