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Study on Fingerprint and in vitro Antiviral Activity of Lonicera japonica Polysaccharide
DING Jie ，YAN Guangling ，YANG Pei ，ZHANG Yongqing ，LIU Yuhong （1. School of Pharmacy，Shandong
1
1
1
1，2
1，2
University of Traditional Chinese Medicine，Jinan 250355，China；2. Fine Breeding Engineering Center of
Chinese Medicinal Materials of Shandong Province，Jinan 250355，China）
ABSTRACT OBJECTIVE：To establish the fingerprint of Lonicera japonica polysaccharide，and to investigate in vitro inhibitory
effect of it on respiratory syndrome virus（RSV）. METHODS：Polysaccharide from L. japonica was prepared by water extraction
and twice alcohol precipitation method. After hydrolysis with trifluoroacetic acid，derivatization with hydroxylamine hydrochloride
and pyridine，the fingerprint was established by GC method. The determination was performed on HP-5 capillary column，and the
detector was flame ionization detector；the temperature of the sample inlet was 250 ℃；the temperature of the detector was 300 ℃
（programmed temperature）；the carrier gas was nitrogen（flow rate of 50 mL/min）；split sampling was adopted（split ratio of
60 ∶ 1）；the sample size was 2.0 μL. Using rhamnose as reference substance，GC fingerprint of 12 batches of L. japonica（S1-S12）
was drawn，and the similarity evaluation was performed with Similarity Evaluation System of TCM Fingerprint（2012 edition）.
Cluster analysis and principal component analysis were conducted by using SPSS 21.0 software. Using ribavirin as positive control，
half effective concentration（EC50 ）and treatment index（TI）as indexes，MTT assay was used to investigate in vitro inhibitory
effect of L. japonica polysaccharide on RSV. RESULTS：There were 12 common peaks in GC fingerprint of 12 batches of L.
japonica. The similarity was greater than or equal to 0.994. Seven common peaks were identified，such as rhamnose，arabinose，
fucose，mannose，glucose，galactose，inositol hexaacetate. According to the cluster analysis，12 batches of samples could be
divided into two categories，i.e. S1，S7，S10 and S11 clustered into one category，and others clustered into one category. In
principal component analysis， the eigen values of 3 principal components were all greater than 1 （5.659， 2.745， 1.724
respectively），and their cumulative contribution rate was 84.400％. The comprehensive score of S12 was the highest，the second
was S5，and the lowest was S11. EC50 of total polysaccharide，80％ alcohol precipitated polysaccharide，50％ alcohol precipitated
polysaccharide and 20％ alcohol precipitated polysaccharide of L. japonica （No. S12） were 0.76， 0.61， 1.03， 3.04 g/L，
respectively；TI were 15.36，18.51，11.69，4.22，respectively. EC50 of 80％ ethanol alcohol precipitated polysaccharide was the
lowest，and its TI was close to that of positive control（20.08）. CONCLUSIONS：Established fingerprint provides reference for the
quality evaluation of L. japonica. L. japonica polysaccharide has a certain inhibitory activity on RSV in vitro，and the 80％ alcohol
precipitated polysaccharide has the strongest activity.
KEYWORDS Lonicera japonica polysaccharide；Fingerprint；Gas chromatography；Principal component analysis；Antivirus
activity

金 银 花 为 忍 冬 科 植 物 忍 冬（Lonicera japonica 金银花多糖对呼吸道合胞病毒（RSV）的体外抑制活性，
Thunb.）的干燥花蕾或带初开的花，始载于《名医别录》， 旨在为金银花临床药效物质基础的进一步阐释提供
为上品，临床上广泛用于治疗多种上呼吸道病毒感染， 参考。
疗效显著 [1-3] 。金银花多糖是该药材发挥降血糖、降血 1 材料
脂、调节免疫、抗肿瘤、抗病毒等功效的重要活性成分之 1.1 仪器
一 [4-7] 。然而，现有金银花的质量评价主要集中在绿原 6980 型 GC 仪（美国 Agilent 公司）；Model 680 型酶
酸、木犀草苷等小分子成分的含量控制上 [1，8] ，而对于多 标仪（美国Bio-Rad公司）；3111型CO2细胞培养箱（美国
糖等重要有效成分的含量却未作规定，使得金银花药材 Thermo Fisher Scientific 公司）；BT25S 型十万分之一电
的质量标准不够完善。指纹图谱是目前应用广泛的质 子天平（德国 Sartorius 公司）；5427R 型高速低温离心机
量评价方法之一，具有整体性、特征性、关联性等特点， （德国 Eppendorf 公司）；YRE-201D 型旋转蒸发仪（巩义
[9]
适用于中药复杂体系的质量研究 。笔者通过检索文献 市予华仪器责任有限公司）。
发现，虽已有金银花指纹图谱的相关研究报道，但仍以 1.2 药材与试剂
小分子成分作为指标，缺乏以金银花多糖等大分子有效 金银花药材购自于山东省漱玉平民大药房、同仁堂
成分为指标的指纹图谱研究。为此，本研究采用气相色 等药房，经山东中医药大学药学院李佳教授鉴定为真
谱法（GC）建立金银花多糖指纹图谱，并进行聚类分析 品。样品信息见表1。
和主成分分析，旨在为金银花药材的质量控制提供依 鼠李糖对照品（批号：C10110722，纯度：≥99％）、阿
据。此外，本课题组前期研究发现，金银花多糖具有抗 拉伯糖对照品（批号：C10127747，纯度：≥99％）、岩藻糖
H1N1流感病毒的作用，主要机制包括调节免疫、促进免 对照品（批号：C10111725，纯度：≥99％）、甘露糖对照品
疫以及抑制炎症反应等 。基于此，本研究初步考察了 （批号：C10122746，纯度：≥99％）、葡萄糖对照品（批号：
[7]

·1062 · China Pharmacy 2020 Vol. 31 No. 9 中国药房 2020年第31卷第9期