高效液相色谱-电化学法测定四季青叶中4种化合物的含量

HPLC-ECD simultaneous determination of four compounds in leaves of Ilex purpurea Hassk.

目的:建立同时测定四季青叶中原儿茶酸、咖啡酸、槲皮素和山奈酚4种成分含量的高效液相色谱-电化学(HPLC-ECD)法。方法:采用 Zorbax SB-C_(18)(150 mm×4.6 mm,5.0 μm)色谱柱;以(A)甲醇、(B)水-0.1%醋酸为流动相,梯度洗脱:0min 时 A-B(20:80),9 min 时 A-B(24:76),30 min 时 A-B(50:50),40 min 时 A-B(65:35);流速为0.8 mL·min^(-1),柱温为30℃,电化学检测器的工作电位为0.7 V,采用标准曲线法对4种化合物进行定量。结果:在选定的色谱条件下原儿茶酸、咖啡酸、槲皮素和山奈酚的检出限分别为3,6,2,5 ng,定量限分别为9.1,20.0,7.2,16.0 ng。4种化合物的平均加样回收率均在95.7%～102.6%之间,RSD 在1.6%～2.9%之间。结论:方法简便、准确,可用于同时检测四季青叶中4种化合物的含量。

Objective： To establish an HPLC - ECD method for simultaneous determination of the contents of protocatechuic acid, caffeic acid, quercetin and kaempferol in leaves of llex purpurea Hassk.. Method： The separation was performed on Zorbax SB - C18 column （ 150 mm × 4. 6 mm,5.0 μm） by gradient elution with （A） methanol - （B） water containing 0. 1% acetic acid [0 min,A - B （20： 80） ;9 min,A - B （24： 76） ;30 min,A - B （50： 50） ; 4-0 min, A - B （65：35 ） ] as the mobile phase at a flow rate of 0. 8 mL min^ - 1. The detection was done at 0. 7 V and 30℃. Calibration curves were applied to the determination of four compounds. Results ：The limits of detection were 3 ng for protocatechuic acid,6 ng for caffeic acid,2 ng for quercetin and 5 ng for kaempferol and the limits of quantification were 9. 1 ng for protocatechuic acid,20. 0 ng for caffeic acid, 7.2 ng for quercetin and 16. 0 ng for kaempferol. The average recoveries of these compounds were between 95.7% - 102. 6% with RSDs of 1.6% - 2. 9%. Conclusion：The method is simple ,rapid ,reproducible and accurate. It can be used for the routine analysis of the herb of Ilex purpurea Hassk..