多波长HPLC法同时测定青黛药材中靛蓝和靛玉红的含量

Simultaneous determination of indigo and indirubin in indigo naturalis by HPLC with multiple UV wavelength detection

目的:利用多波长高效液相色谱法,建立同时测定青黛药材中2种有效成分(靛蓝和靛玉红)含量的方法,并对其质量特征进行分析。方法:采用HPLC法,Phenomenex C18色谱柱(250 mm×4.6 mm,5μm),柱温30℃;以甲醇-水溶液(70∶30)为流动相洗脱,流速1.0 ml/min;检测波长286 nm(检测靛蓝)、292 nm(检测靛玉红),同时测定不同产地青黛药材中靛蓝和靛玉红含量。结果:在20 min内青黛中靛蓝和靛玉红2种有效成分被完全分离。靛蓝在5.19~83.04μg/ml范围内线性关系良好(r=0.999 9),平均回收率为100.46%,RSD为2.18%;靛玉红在0.364~5.825μg/ml范围内线性关系良好(r=0.999 9),平均回收率为101.57%,RSD为2.22%。结论:本方法合理、便捷、快速简便、准确、重复性好,能同时测定青黛中靛蓝和靛玉红含量,符合现行《中国药典》简便、快速、准确的要求。

Objective： To establish an HPLC method with multiple UV wavelengths detection to simultaneously determine the contents of indigo and indirubin in indigo naturalis and to characterize the quality of indigo naturalis. Methods ： The samples were separated on a Phenomenex column （ 250 mm× 4.6 mm, 5 μm）. The column temperature was set at 30 ℃. The mobile phase was methanol - water （70： 30） and the flow rate was 1.0 ml/min. A multiple detector was used for determination of indigo at 286 nm and of indirubin at 292 nm. Result： The complete separation was obtained within 20 rain for the two compounds （ indigo and indirubin）. The linear correlation was satisfied in the range from 5.19 to 83.04 μg/ml （ r = 0.999 9）for indigo and from 0.364 to 5. 825 μg/ml （ r = 0. 999 9 ） for indirubin. The average recoveries of the low, middle and high concentrations were 98.78%, 100.44%, 104.35%, respectively for indigo; 96.20%, 98.63%, 104.03% respectively for indirubin. Twenty - seven batches of samples were divided into three categories, which reflected the inherent quality characteristics in different regions of indigo naturails. Conclusion： Compared with the method specified in Vol I Chinese Pharmacpeia 2015, the method is simple and the result is reliable with good repeatability and can be used for the simultaneous determination of indigo and indirubin.