Study on the Mixed-Mode HPLC Separation of Shuang-Huang-Lian and Xue‑Bi‑Jing Injections Based on an Ionic Liquid Column

Abstract

Both Shuang-Huang-Lian and Xue-Bi-Jing injections are clinically proven efficient and low cost medicines. However, the studies of their chromatographic separation are limited. In this study, a column based on ionic liquid stationary phase was applied for the first time for the analysis of these two injections. For Shuang-Huang-Lian injection, U-shaped plots of retention factors vs. mobile phase constitutions were obtained. This unique kind of separation behavior allowed a flexible adjustment of retention by tuning the mobile phase constituents. Then acids group was analyzed with hydrophilic chromatography mode, while glycosides group was analyzed with reversed phase chromatography mode. Similarly, constituents in Xue-Bi-Jing injection were analyzed with mixed mode high performance liquid chromatography (HPLC). Higher, medium and lower polarity groups were analyzed with hydrophilic interaction, reversed phase and normal phase liquid chromatography modes, respectively. Results from this work provided valuable information for the separation of complicated systems such as traditional Chinese medicine injections.Keywords: ionic liquid stationary phase, mixed-mode chromatography, Shuang-Huang-Lian injection, Xue-Bi-Jing injection IntroductionThe formulations of traditional Chinese medicines (TCMs) are becoming increasingly popular worldwide, mainly because of their high efficiency and low cost. However, TCM formulations are usually based on a whole plant and/or combination of several herbs, which contain more than dozens to hundreds of components. 1 Therefore, in order to ensure both the safety and efficacy of TCM formulations, it is necessary to establish simple and effective methods to determine as many constituents as possible.2,3 Liquid chromatography (LC) has been applied as a rapid and economic method for the determination of multi-component in TCMs. 4 Among the liquid chromatography analytical methods, high performance liquid chromatography (HPLC) is the most popular one, due to its easy operation, wide suitability and high accuracy for the qualitative and quantitative analysis of TCMs. [5][6][7] However, in practice, some factors, such as efficiency, selectivity and sensitivity have been found to affect the analysis of complicated systems with HPLC. 8Chromatographic stationary phase is the core of HPLC system. Therefore, the development of novel stationary phases with high separation efficiency and excellent selectivity has always been the focus point of improving the separating ability of HPLC. Accordingly, in this work novel ionic liquid (IL) bonded stationary phase has been studied. IL is a kind of novel material composed of ions and at the same time remains in liquid state at a temperature below 100 °C.9 ILs have demonstrated the versatility and tuneability in separation science. 10 Many types of ILs were immobilized on a support like silica or monolith as stationary phases for liquid chromatography. 11,12 Due to the designable character of the IL molecule, multi interactions between the IL stationary phas...