Integrated strategy of derivatization and separation for sensitive analysis of salvianolic acids using capillary electrophoresis with laser-induced fluorescence detection

Abstract

Salvianolic acids (SAs) are a class of natural active substances that have been widely used in clinical treatment and food adjuvant therapy. In this work, we found that SAs could form the ternary complex with borax and 2-hydroxypropyl-β-cyclodextrin (HP-β-CD), thereby sensitizing their weak intrinsic fluorescence to maximized 92-fold for salvianolic acid B (SAB). The formation of ternary complex was dynamic and could complete once mixed, and the fluorescence intensity remained stable within 3 h. On this basis, an integrated strategy of derivatization and separation was proposed for the sensitive analysis of SAs using capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection. After the systematic investigation, a solution consisting of 20 mM borax and 20 mM HP-β-CD (pH 8.5) was selected as the running buffer. By the direct injection of SAs, the dynamic derivatization was realized in the capillary, and the baseline separation was achieved within 6 min, with the detection limits of 1.2-21.2 nM for four SAs (S/N=3). Then, the developed CE-LIF method was successfully applied to the quantitative analysis of SAs in four traditional ginsengs, including Salvia miltiorrhiza, Codonopsis pilosula, Panax quinquefolius and Panax ginseng with the recoveries ranging from 95.2% to 110.7%. Except for four target SAs, a large number of unknown electrophoretic peaks had also been observed in four ginsengs, and then were utilized for the identification of ginseng species via principal component analysis. Furthermore, a hypoxia/reoxygenation model was constructed using Rat cardiomyocyte H9c2 cells, and subsequently, the developed method was applied to continuously monitor the consumption of SAB in cell culture medium after its intervention.