UPLC-MS/MS法测定不同产地浮萍中9种核苷类成分的量

Determination of nine kinds of nucleosides in Spirodelae Herba from different habitats by UPLC-MS/MS

目的建立超高效液相色谱三重四级杆串联质谱法（UPLC-MS/MS）同时测定浮萍中9种核苷类成分的方法,分析比较不同产地样品中9种成分的差异。方法采用Waters XbridgeTM Amide色谱柱（150 mm×4.6 mm,3.5μm）;以0.1%甲酸的水（A）-0.1%甲酸的乙腈（B）溶液为流动相,梯度洗脱,体积流量为0.5 m L/min;柱温30℃,采用电喷雾离子源,正离子检测方式,得到相应的提取离子流图,以峰面积进行定量,用主成分分析和聚类分析方法进行综合评价。结果 9种核苷类成分具有良好的线性关系,r值均大于0.998 9,方法精密度、重复性和稳定性的RSD值均小于3.5%,加样回收率在94.4%~101.9%,RSD在1.73%~3.58%。不同产地浮萍药材中9种核苷在组成和量上均有差异,其中2′-脱氧尿苷、2′-脱氧肌苷在13个产地样品中量均较低,以胞嘧啶最低;尿苷、肌苷、黄嘌呤量较高;13个产地中安徽黄山浮萍中9种核苷总量最高。结论该方法操作简便、准确、重复性好,可为浮萍药材的质量标准制定提供依据。

Objective To establish a UPLC-MS/MS method for the simultaneous determination of nine kinds of nucleosides in Spirodelae Herba, and analyze and investigate the difference of nucleosides ingredients in samples from 13 habitats. Methods The determination was performed on the Waters XbridgeTM Amide-C18（150 mm × 4.6 mm, 3.5 μm）, the 0.1% formic acid-water（A） and 0.1% acetonitrile-water（B） in gradient elution as mobile phase. The flow rate was 0.5 m L/min. The column temperature was 30 ℃. MS instrument was equipped with ESI＋ ion source. Gaining the extracted ion chromatograms, then the peak area for quantitative, principal component analysis（PCA）, and hierarchical clustering analysis（HCA） were used to comprehensive evaluation. Results All nine kinds of nucleoside showed good linearity（r ≥ 0.998 9）. The RSD of the precision, repeatability, and stability tests were less than 3.5%. The average recovery rates were in the range of 94.4%—101.9%, RSD were in the range of 1.73%- 3.58%. There are differences in the composition and content from different habitats. The content of 2′-deoxyuridine and 2′-deoxyinosine in the lower levels, cytosine was the lowest, while the contents of uridine, inosine, and xanthine were in higher levels. The largest content was in Spirodelae Herba from Huangshan, Anhui province. Conclusion The method for the simultaneous determination of the nine kinds of nucleosides is simple, accurate, and reproducible, that will provide the basis for the formulation of herbs Spirodelae Herba for quality control standards.