金银花多糖的指纹图谱及体外抗病毒活性研究
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篇名: 金银花多糖的指纹图谱及体外抗病毒活性研究
TITLE: Study on Fingerprint and in vitro Antiviral Activity of Lonicera japonica Polysaccharide
摘要: 目的:建立金银花多糖的指纹图谱,并考察其对呼吸道合胞病毒(RSV)的体外抑制作用。方法:通过水提、二次醇沉制备金银花多糖;所得多糖经三氟乙酸水解和盐酸羟胺、吡啶等衍生化后,采用气相色谱法(GC)建立指纹图谱。色谱柱为HP-5毛细管色谱柱,检测器为火焰离子化检测器,进样口温度为250℃,检测器温度为300℃,程序升温,载气为氮气(流速50mL/min),分流进样(分流比60∶1),进样量为2.0μL。以鼠李糖为参照,绘制12批金银花药材样品(S1~S12)的GC指纹图谱,采用《中药色谱指纹图谱相似度评价系统(2012版)》进行相似度评价并确定共有峰;采用SPSS21.0软件进行聚类分析和主成分分析。以利巴韦林为阳性对照,以半数有效浓度(EC50)、治疗指数(TI)为指标,采用MTT法考察金银花多糖对RSV的体外抑制作用。结果:12批金银花药材样品的GC指纹图谱有12个共有峰,相似度≥0.994;共指认鼠李糖、阿拉伯糖、岩藻糖、甘露糖、葡萄糖、半乳糖、肌醇六乙酸酯等7个共有峰。12批药材样品共聚为两类,即S1、S7、S10、S11聚为一类,其余聚为一类;主成分分析中,3个主成分的特征值均大于1(分别为5.659、2.745、1.724),其累积贡献率为84.400%。样品综合得分以S12最高,S5次之,S11最低。金银花(S12)总多糖、80%醇沉多糖、50%醇沉多糖、20%醇沉多糖的EC50值分别为0.76、0.61、1.03、3.04g/L,TI值分别为15.36、18.51、11.69、4.22;其中,80%醇沉多糖的EC50值最低,TI值与阳性对照(20.08)接近。结论:所建指纹图谱可为金银花药材的质量评价提供参考。金银花多糖对RSV具有一定的体外抑制活性,且以80%醇沉多糖活性最强。
ABSTRACT: OBJECTIVE:To establis h 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. S 1,S7,S10 and S 11 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 S 12 was the highest ,the second was S 5,and the lowest was S 11. EC 50 of total polysaccharide ,80% alcohol precipitated polysaccharide ,50% alcohol precipitated polysaccharide and 20% alcohol precipitated polysaccharide of L. japonica (No. S 12) were 0.76,0.61,1.03,3.04 g/L, respectively;TI were 15.36,18.51,11.69,4.22,respectively. EC 50 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.
期刊: 2020年第31卷第09期
作者: 丁洁,闫光玲,杨培,张永清,刘玉红
AUTHORS: DING Jie,YAN Guangling,YANG Pei,ZHANG Yongqing,LIU Yuhong
关键字: 金银花多糖;指纹图谱;气相色谱;主成分分析;抗病毒活性
KEYWORDS: Lonicera japonica polysaccharide;Fingerprint;Gas chromatography ;Principal component analysis ;Antivirus
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