Screening of active ingredients from traditional Chinese medicine using two-dimensional liquid chromatography-mass spectrometry: A case study of screening neuraminidase inhibitors from Schizonepetae spica

The traditional methods of screening active ingredients from traditional Chinese medicine (TCM) are primarily based on chemical separation followed by activity evaluation, which involve long experimental cycles and large workloads. However, identifying active ingredients in TCM based on the targets related to a disease results in significantly improved screening efficiency. In this study, we developed a method for screening active components using two-dimensional liquid chromatography coupled with mass spectrometry (2D LC-MS). Briefly, the TCM extract was co-incubated with the target to form target-ligand complexes. Then, the incubation solution was loaded onto a 1D size-exclusion chromatography column, where the target-ligand complexes were separated from the unbound free compounds. Subsequently, a 2D reversed-phase chromatography column was used to dissociate the target-ligand complexes. Chromatographic separation was further optimized to ensure separation of the released ligands and inactive target. Finally, the dissociated ligands were analyzed using high-resolution mass spectrometry for structural identification. As an example, we screened the neuraminidase inhibitors from Schizonepetae spica using the method developed in this study and successfully identified a total of 40 ligands. The proposed 2D LC-MS method can be used for screening Other active ingredients in TCM, and the screening model for neuraminidase inhibitors established in this study can be used for screening neuraminidase inhibitors from Other TCM.

LC-MS; Neuraminidase inhibitor; Schizonepetae spica; Two-dimensional liquid chromatography.