不同产地水冬瓜叶HPLC指纹图谱建立、化学模式识别分析及含量测定

Establishment of HPLC Fingerprint,Chemical Pattern Recognition Analysis and Content Determination of the Leaves of Toricellia angulata from Different Regions

目的:为水冬瓜叶药材的质量控制提供参考。方法:采用高效液相色谱法(HPLC),以Agela Promosil C18为色谱柱,以0.2%磷酸水溶液-乙腈(梯度洗脱)为流动相,流速为1.0 mL/min,检测波长为210 nm,柱温为35℃,进样量为10μL。采用《中药色谱指纹图谱相似度评价系统(2004 A版)》建立10批不同产地水冬瓜叶的HPLC指纹图谱并进行相似度评价,通过与对照品色谱图比对进行色谱峰的指认。采用聚类分析、主成分分析(PCA)法和偏最小二乘法-判别分析(PLS-DA)法进行化学模式识别分析,并筛选质量差异标志物。采用相同HPLC法对水冬瓜叶中金丝桃苷和异槲皮苷的含量进行测定。结果:10批不同产地水冬瓜叶HPLC指纹图谱与对照指纹图谱的相似度为0.923~0.983。共确定了11个共有色谱峰,并指认出了峰4、5分别为金丝桃苷、异槲皮苷。聚类分析和PCA、PLS-DA结果均显示,10批水冬瓜叶可分为2类,其中,Y10聚为一类、其余聚为一类;经PLS-DA分析,筛选出了6个变量投影重要性值大于1的共有峰(峰4、3、10、2、6、11)。10批样品中金丝桃苷和异槲皮苷的平均含量分别为0.47~6.97、0.21~1.87 mg/g。结论:所建立的HPLC指纹图谱和含量测定方法稳定、可靠,初步筛选出了不同产地水冬瓜叶中金丝桃苷等6个质量差异性标志物,可用于不同产地水冬瓜叶药材的质量控制。

OBJECTIVE:To provide reference for the quality control of the leaves of Toricellia angulata.METHODS:HPLC method was adopted.The determination was performed on Agela Promosil C18 column with 0.2%phosphoric acid solution-acetonitrile(gradient elution)as mobile phase at the flow rate of 1.0 mL/min.The detection wavelength was set at 210 nm,and column temperature was 35℃.The sample size was 10μL.HPLC fingerprint of 10 batches of the leaves of T.angulata was established and similarity evaluation was conducted by using Similarity Evaluation System of TCM Chromatographic Fingerprint(2004 edition).The chromatographic peak was identified by comparing with the chromatogram of reference substance.Cluster analysis,PCA and PLS-DA were used to identify chemical patterns,and the quality differential markers were screened.The contents of hyperoside and isoquercitrin were determined by the same HPLC.RESULTS:The similarities of HPLC fingerprint of 10 batches of the leaves of T.angulata with control fingerprint were 0.923-0.983.A total of 11 common peaks were identified,and the peaks 4 and 5 were hyperoside and isoquercitrin,respectively.Results of cluster analysis,PCA and PLS-DA showed that 10 batches of leaves of T.angulata could be divided into two categories,Y10 was clustered into one category,and others were clustered into one category.PLS-DA analysis showed that 6 common peaks(peaks 4,3,10,2,6 and 11)with variable importance projection(VIP)greater than 1 were selected.Average contents of hyperoside and isoquercitrin in 10 batches of the leaves of T.angulata were 0.47-6.97,0.21-1.87 mg/g,respectively.CONCLUSIONS:Established HPLC fingerprint and the method for content determination are stable and reliable,and can be used for the quality control of the leaves of T.angulata from different areas.Six quality differential markers including hyperoside in the leaves of T.angulata from different areas are preliminarily screened.