HPLC法同时测定口腔溃疡散II号中靛蓝和靛玉红的含量

目的：建立同时测定口腔溃疡散II号中靛蓝和靛玉红含量的HPLC方法，用以检测该制剂的质量。方法：采用正交设计优化超声提取方法，以溶剂体积、提取时间、提取温度为主要影响因素，以靛蓝和靛玉红的提取总量为评价指标。采用ZORBAX SB-C₁₈（4.6 mm×250 mm，5 μm）色谱柱，流动相组成为乙腈-水（65：35），流速1.0 mL·min^-1，检测波长286 nm，柱温20 ℃。结果：确定最佳提取方法为0.3 g供试品用30 mL N，N-二甲基甲酰胺在40~45 ℃条件下超声提取40 min。靛蓝和靛玉红进样量分别在59.77~1 195 ng（r=1.000）和6.300~126.0 ng（r=0.999 8）范围内与色谱峰面积呈良好的线性关系；平均回收率分别为99.1% 和100.4%，RSD分别为2.8%和2.1%（n=6）。6批样品中靛蓝、靛玉红的含量分别为6.219~6.423、0.495~0.631 mg·g^-1。结论：本文建立的方法经方法验证，可用于口腔溃疡散II号中靛蓝和靛玉红的定量分析。

Objective: To establish an HPLC method for simultaneous determination of indigo and indirubin in Kouqiangkuiyang powders II, so as to control the quality of this preparation. Methods: Orthogonal array design was used to optimize the extracting process. The main influential factors of extracting were solvent volume, extracting time and extracting temperature. The conditions of the extracting were evaluated by the total content of indigo and indirubin. The sample was separated by a ZORBAX SB-C₁₈(4.6 mm×250 mm, 5 μm)column with the mobile phase consisting of acetonitrile-water(65:35)at the flow rate of 1.0 mL·min^-1, UV detection wavelength at 286 nm and the column temperature at 20 ℃. Results: The analysis indicated that the optimum conditions for indigo and indirubin from Kouqiangkuiyang powders II were as follows: the sample(0.3 g)was extracted with 30 mL N, N-dimethyl formamide under ultrasonic extractor of 40-45 ℃ for 40 min. Indigo and indirubin had good linearity in the ranges of 59.77-1 195 ng(r=1.000)and 6.300-126.0 ng(r=0.999 8). The average recovery and RSD were 99.1%, 100.4% and 2.8%, 2.1% respectively(n=6).The content of indigo and indirubin in six samples was 6.219-6.423 mg·g^-1、0.495-0.631 mg·g^-1. Conclusion: The method is simple, reliable, specific, repeatable and could be applied to quantitative determination of indigo and indirubin in Kouqiangkuiyang powders II.