HPLC-MS/MS法同时测定玄参中8个成分的含量

Simultaneous Determination of Eight Components in Radix Scrophulariae by HPLC-MS/MS

目的:建立HPLC-MS/MS法同时测定玄参中梓醇、哈巴苷、栀子苷、类叶升麻苷、安格洛苷C、肉桂酸、哈巴俄苷、5-羟甲基糠醛8个成分的含量。方法:采用Agilent ZORBAX Eclipse XDB-C18色谱柱(150 mm×4.6 mm,5μm),柱温为30℃,以甲醇-0.05%甲酸水为流动相,梯度洗脱,流速为0.6 m L·min-1,进样量2μL,ESI源,正离子-MRM模式下检测;结果:梓醇、哈巴苷、栀子苷、类叶升麻苷、安格洛苷C、肉桂酸、哈巴俄苷、5-羟甲基糠醛质量浓度分别在0.194 0~7.76(r=0.999 0)、0.173 8~32.85(r=0.999 8)、0.394~23.64(r=0.996 4)、0.279 5~16.77(r=0.997 4)、0.3095~12.38(r=0.999 4)、0.087~5.220(r=0.999 1)、0.264 5~31.74(r=0.999 8)、0.034 3~1.370 0μg·mL^(-1)(r=0.999 2)线性关系良好,检出限(LOD)为5~50 ng·mL^(-1),平均回收率为91.33%~99.78%。样品测定结果显示,玄参中哈巴俄苷、哈巴苷及安格洛苷C含量较高,分别为1.517~9.735、0.289~5.997、0.898~3.568 mg·g-1。不同来源的样品中,8个成分的含量及各个成分之间比例存在较大差异。结论:本实验建立了在22 min内同时测定玄参中8个成分含量的方法,灵敏度高,准确度好,实验结果可为玄参质量提供参考。

Objective： To establish a method for simultaneous determination of eight components in Radix Scrophu- lariae including catalpol, harpagoside, geniposide, acteoside, angoroside C, cinnamic acid, harpagide, 5-HMF. Methods： Chromatographic separation was performed on an Agilent ZORBAX Eclipse XDB-C18 column （ 150 mm × 4. 6 mm, 5 Fro） at 30 ℃ of column temperature. 0. 05% aqueous formic acid （A） and methanol （B） were adopted as mobile phase with a gradi- ent elution. The flow rate was set at 0. 6 mL min-1. The injection volume was 2 μL. Detection was carried out on a triple qua- drupole mass spectrometer in the positive ion mode using an electrospray source. Multiple reaction monitoring（MRM） mode was employed. Results： The developed method showed good linearity in the range of O. 194 0-7. 76 μg mL-l （r = O. 999 0 ） for catalpol, 0. 173 8-32. 85 μg mL- 1 （ r = 0. 999 8 ） for harpagoside, 0. 394-23.64 μg mL-1 （ r = 0. 996 4 ） for geniposide, 0. 279 5-16. 77 μg mL-1 （ r = 0. 9974 ） for acteoside, 0. 309 5-12. 38μg mL- 1（ r = 0. 999 4 ） for angoroside C, 0. 087- 5. 220 μg mL- 1（ r = 0. 9991 ） for cinnamic acid, 0. 264 5-31.74 μg mL- 1 （ r =0. 999 8 ） for harpagide, 0. 034 3- 1. 370 0 μg mL- 1 （ r = 0. 999 2） for 5-HMF, respectively. The limit of detection was 5-50 ng mL- 1. The recoveries varied between 91.33% -99.78%. Harpagide, harpagoside and angoroside C were found to be the most abundant components in Radix Scrophulariae. There were significant differences in distribution of these components among the samples from different origins. Conclusion： The established method of determination of 8 kinds of component content in Radix Scrophulariae within 22 min is proved to be accurate and sensitive. The experimental results can provide reference for the quality of Radix Scrophulariae.