水蔓菁中5种主要化学成分的分离及含量测定

Separation and Content Determination of Five Main Chemical Constituents from Pseudolysimachion linariifolium subsp.dilatatum

目的分析水蔓菁中的苷类化合物,建立同时测定5种主要苷类化合物含量的方法。方法对水蔓菁的苷类化合物进行分离和纯化,根据化合物的波谱数据进行结构鉴定;运用高效液相色谱(HPLC)的面积归一化法测定其5种主要苷类化合物的含量。采用安捷伦Extend-C_(18)色谱柱(250 mm×4.6 mm,5.0μm);甲醇(A)-0.1%甲酸水溶液(B)为流动相,梯度洗脱;流速:1.0 mL·min^(-1);柱温:25℃;检测波长:260、340 nm。结果分离鉴定了水蔓菁中5个主要苷类化合物,分别为毛蕊花苷(1)、梓苷(2)、胡黄连苷Ⅱ(3)、梓醇6-咖啡酸酯(4)、水蔓菁苷Ⅱ(5),纯度分别为99.5%、99.5%、99.8%、99.8%及98.1%。在75 min内,水蔓菁中毛蕊花苷、梓苷、胡黄连苷Ⅱ、梓醇6-咖啡酸酯和水蔓菁苷Ⅱ能达到良好的分离,分别在0.597~9.552,0.212~3.398,0.200~3.146,0.300~4.806,0.300~4.813μg范围内呈良好线性关系(r分别为0.9999、0.9999、0.9999、0.9998、0.9999),平均回收率为98.57%、103.10%、103.97%、99.66%、103.60%,相对标准偏差(RSD)分别为2.45%、1.83%、2.11%、3.32%、1.96%。结论该研究建立的方法操作简便、准确,专属性强,重复性好,可用于水蔓菁中5种主要苷类化合物的含量测定,并为水蔓菁药材及其制剂的质量控制提供依据。

Objective To analyze the glycosides from Pseudolysimachion linariifolium subsp.dilatatum,and establish a method for simultaneous determination of five main glycosides.Methods The glycosides of P.linariifolium subsp.dilatatum were isolated and purified by means of various of chromatography materials and separation technologies,and their structures were established through spectroscopic methods.The purities of five main glycosides were determined by HPLC area normalization method,and the analysis was performed at 25℃on an Extend-C_(18) column(250 mm×4.6 mm,5.0μm),eluted with a gradient program using methanol(A)-0.1%formic acid aqueous solution(B)as the mobile phase.The flow rate was 1.0 mL·min^(-1),and the detection wavelengths were 260 nm and 340 nm.Results Five compounds were isolated and identified,included verproside(1),catalposide(2),pictosideⅡ(3),verminoside(4)and linariifoliosideⅡ(5),with the purities of 99.5%,99.5%,99.8%,99.5%,98.1%,respectively.The five components,verproside(1),catalposide(2),pictosideⅡ(3),verminoside(4),and linariifoliosideⅡ(5)could be well separated within 75 minutes.Their calibration curves were with good linear relationships in the ranges of 0.597-9.552μg(r=0.9999),0.212-3.398μg(r=0.9999),0.200-3.146μg(r=0.9999),0.300-4.806μg(r=0.9998),0.300-4.813μg(r=0.9999),respectively.The average recovery rates were 98.57%,103.10%,103.97%,99.66%,103.60%,and RSDs were 2.45%,1.83%,2.11%,3.32%,1.96%.Conclusion The method established in this research for simultaneous determination of the five components was specific,simple,accurate,and reproducible,which can provide a scientific basis for quality control of P.linariifolium subsp.dilatatum.