HPLC法测定舞阳贝母水溶性成份含量及指纹图谱分析

Determination of Water-Soluble Components and Fingerprint Analysis of Fritillaria Wuyangensis Bulbus by HPLC

目的:建立舞阳贝母中9种核苷类成份的含量测定方法,初步构建HPLC指纹图谱,为其质量控制提供参考。方法:采用YMC-Triart C18色谱柱(250 mm×4.6 mm,5μm),流动相为甲醇-水,梯度洗脱,流速1.0 ml/min,检测波长260 nm,柱温30℃。对9批舞阳贝母样品进样分析,测定9种核苷类成份的含量,并采用“中药色谱指纹图谱相似度评价系统(国家药典委员会,2012版)”进行相似度评价。结果:定量分析方法学考察结果良好,9个核苷类成份加样回收率96.74%~101.90%;9批样品中尿嘧啶、胞苷、次黄嘌呤、尿苷、腺嘌呤、鸟苷、胸苷、腺苷、2′-脱氧腺苷的含量分别为3.92~63.74、8.72~109.23、4.16~21.77、135.08~717.54、5.42~67.65、318.39~561.04、114.33~269.32、280.20~464.09、51.81~171.30μg/g。初步构建舞阳贝母水溶性成份HPLC指纹图谱,9批样品相似度在0.822~0.910,共得到15个共有色谱峰,通过保留时间定位确认其中9个色谱峰归属,即峰1为尿嘧啶、峰2为胞苷、峰4为次黄嘌呤、峰5为尿苷、峰8为腺嘌呤、峰9为鸟苷、峰11为胸苷、峰13为腺苷、峰14为2′-脱氧腺苷。结论:建立的9种核苷类成份的含量测定方法简便稳定、重复性好,结合HPLC指纹图谱分析能够更加全面地反映舞阳贝母的质量,为其深度开发提供了研究基础。

Objective:A method for the determination of 9 nucleoside components in Fritillaria Wuyangensis Bulbus was established,and the high performance liquid chromatography(HPLC)fingerprint was preliminarily constructed to provide a reference for its quality control.Methods:The analysis was performed on a YMC-Triart C18 column(250 mm×4.6 mm,5μm)with a gradient of methanol-water at a flow rate of 1.0 ml/min,the detective wavelength was set at 260 nm,the column temperature was set at 30℃.Nine batches of Fritillaria Wuyangensis Bulbus samples were injected and analyzed,the contents of nine nucleosides were determined,and the similarity was evaluated by the“similarity evaluation system of chromatographic fingerprint of traditional Chinese medicine(2012 edition,National Pharmacopoeia Committee)”.Results:The results of quantitative analysis were good,and the recovery rate of nine components was 96.74%~101.90%.The content of uracil,cytidine,hypoxanthine,uridine,adenine,guanosine,thymidine,adenosine and 2′-deoxyadenosine in 9 batches of samples were in the range of 3.92~63.74,8.72~109.23,4.16~21.77,135.08~717.54,5.42~67.65,318.39~561.04,114.33~269.32,280.20~464.09,51.81~171.30μg/g,respectively.HPLC fingerprint of water-soluble components of Fritillaria Wuyangensis Bulbus was preliminarily established.The similarity of 9 batches of Fritillaria Wuyangensis Bulbus was between 0.822 and 0.910,and a total of 15 common peaks were identified.Nine chromatographic peaks were identified by retention time localization.Peak 1 was uracil,peak 2 was cytidine,peak 4 was hypoxanthine,peak 5 was uridine,peak 8 was adenine,peak 9 was guanosine,peak 11 was thymidine,peak 13 was adenosine,and peak 14 was 2′-deoxyadenosine.Conclusion:The established method for the determination of 9 nucleosides is simple,stable and reproducible.Combined with HPLC fingerprint analysis,it can reflect the quality of Fritillaria Wuyangensis Bulbus more comprehensively,providing a research basis for its in-depth development.