Simultaneous Determination of Aristoloxazines, Aristolochic Acids, and Aristolactams Using HPLC–Fluorescence Detection with a Post-column Microreactor: Application in Identifying New Aristoloxazines

Aristoloxazines (AXs), aristolochic acids (AAs), and aristolactams (ALs) are potent genotoxins found in Asarum and Aristolochia plants, many of which are commonly used as herbal medicines. Emerging evidence indicates that these compounds contaminate arable soil during the cultivation of Aristolochiaceae herbs. Currently, no method exists for their simultaneous detection. In this study, we developed a high-performance liquid chromatography (HPLC)-based method for their determination. This method employs an iron powder-packed microreactor to convert non-fluorescent AXs and AAs into naturally fluorescent ALs, enabling fluorescence detection after HPLC separation. After being validated against LC–MS/MS analysis, the method was applied to quantify these genotoxins in herbal and soil samples, detecting AXs at concentrations as high as hundreds of μg/g in some Asarum samples. Given that Asarum plants are widely used in herbal medicine, these results reveal previously unrecognized human exposure to genotoxins that warrants the attention of both the general public and regulatory agencies.