猪苓多糖的PMP柱前衍生化-HPLC指纹图谱研究
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篇名: 猪苓多糖的PMP柱前衍生化-HPLC指纹图谱研究
TITLE: Study on PMP Pre-column Derivatization-HPLC Fingerprint of Polyporus Polysaccharide
摘要: 目的:建立猪苓多糖的柱前衍生化-高效液相色谱(HPLC)指纹图谱,并测定其主要单糖组分含量,为猪苓药材的质量评价提供参考。方法:11批不同产地猪苓药材经水提醇沉、Sevage除蛋白后得到猪苓多糖。多糖经三氟乙酸(TFA)水解和1-苯基-3-甲基-5-吡唑啉酮(PMP)衍生化后,进行HPLC分析。色谱柱为HypersiLBDSC18,流动相为0.1mol/L磷酸盐缓冲液(pH6.84)-乙腈(84∶16,V/V),检测波长为254nm,柱温为30℃,流速为1mL/min,进样量为20µL。采用《中药色谱指纹图谱相似度评价系统》(2012A版)建立11批多糖样品的指纹图谱并进行相似度评价,通过与对照品比对进行色谱峰指认,再采用SPSS23.0软件进行聚类分析,并测定多糖中主要单糖组分的含量。结果:11批猪苓多糖样品HPLC指纹图谱中共呈现出3个共有峰,分别指认为甘露糖、葡萄糖和半乳糖,各批次样品的相似度均大于0.94。聚类分析将11批多糖样品分为3类,药材编号为S1~S6、S8的样品聚为一类,药材编号为S7、S10、S11的样品聚为一类,药材编号为S9的样品单独聚为一类。含量测定结果显示,11批样品中甘露糖含量为1.571~8.771mg/g、葡萄糖含量为26.072~132.194mg/g、半乳糖含量为3.420~36.593mg/g。结论:本研究建立的柱前衍生化-HPLC指纹图谱方法可为猪苓的药材质量评价提供参考;不同批次猪苓多糖中单糖组成相同,指纹图谱特征峰与药材的产地无明显相关性,药材之间的单糖含量存在明显差异。
ABSTRACT: OBJECTIVE:To establish pre -column derivatization-HPLC fingerprint of Polyporus polysaccharide ,and to determine the contents of main monosaccharide components ,so as to provide reference for quality evaluation of Polyporus umbellatus. METHODS :Polyporus polysaccharide was extracted with boiling water and precipitated by ethanol and deproteinized by Sevage from 11 batches of P. umbellatus from different producing areas. The samples were firstly hydrolyzed with trifluoro-acetic acid (TFA)and then derivatized by 1-phenyl-3-methyl-5-pyrazolone(PMP). HPLC analysis was then conducted. The determination was carried out on HypersiL BDS C 18 column with mobile phase composed of 0.1 mol/L phosphate buffer (pH 6.84)-acetonitrile(84∶16,V/V)by gradient elution at the flow rate of 1.0 mL/min. The detection wavelength was set at 254 nm, and column temperature was 30 ℃. The sample size was 20 µL. The similarity of 11 batches of Polyporus polysaccharide was evaluated by using TCM Chromatographic Fingerprint Similarity Evaluation System (2012A edition ),and the contents of main monosassharide components were detected. The peak was identified by comparing with the reference substance ,and cluster analysis was performed by using SPSS 23.0 software. RESULTS :In HPLC fingerprints of the 11 batches of samples ,3 common peaks were identified ,namely mannose ,glucose and galactose. The similarity of all samples was above 0.94. Cluster analysis classified 11 batches of samples into three categories. S 1-S6,and S 8 were grouped into category 1;S7,S10 and S 11 were grouped into category 2;S9 was individually grouped into one category. Results of content determination showed that the contents of mannose ranged from 1.571 to 8.771 mg/g;those of glucose ranged from 26.072 to 132.194 mg/g,and those of galactose ranged from 3.420 to 36.593 mg/g. CONCLUSIONS :Established pre-column derivatization HPLC fingerprints can provide reference for quality evaluation of P. umbellatus . The monosaccharide composition of different batches of Polyporus polysaccharide is the same ;there is no significant correlation between fingerprint characteristic peak and the origin of herbs ;there is significant difference in the content of monosaccharide of P. umbellatus .
期刊: 2020年第31卷第07期
作者: 赖戈娜,贾文玉,罗思婉,周昌园,黎雄,张娴,曾星
AUTHORS: LAI Gena,JIA Wenyu,LUO Siwan,ZHOU Changyuan,LI Xiong,ZHANG Xian,ZENG Xing
关键字: 猪苓多糖;单糖;1-苯基-3-甲基-5-吡唑啉酮;柱前衍生化;高效液相色谱法;指纹图谱
KEYWORDS: Polyporus polysaccharide ;Monosaccharide;PMP;Pre-column derivatization ;HPLC;Fingerprint
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