Chemical heterogeneity of two <i>Bupleurum</i> Radix species based on multiple mass spectrometry techniques

In this study, the chemical profiles and spatial distributions of Bupleurum chinense DC (BC) and Bupleurum scorzonerifolium Willd. (BS) were systematically analyzed by integrating cryosectioning, histochemical localization, matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), inductively coupled plasma-mass spectrometry (ICP-MS), and liquid chromatography mass spectrometry (LC-MS). Histochemical staining indicated that BS contained relatively high levels of total saponins, whereas MALDI-MSI and LC-MS revealed that BC resulted in significantly high levels of characteristic saikosaponins, which exhibited a ring-like enrichment pattern in the phloem. MALDI-MSI elucidated the spatial distribution of multiple bioactive components in Bupleurum roots. Uniform manifold approximation and projection (UMAP) algorithm-based spatial segmentation demonstrated distinct chemical zonation in BC, with a concentric metabolite distribution, whereas BS exhibited asymmetric lateral heterogeneity. ICP-MS analysis revealed K and Ca to be the predominant mineral elements, revealing significant geographical enrichment patterns (e.g., higher K levels in Gansu samples and higher Ca levels in Shanxi samples). Additionally, eight elements (including Fe, Ni, and Cr) displayed significant interspecies differences ( p &lt; 0.05). By combining multiple mass spectrometry techniques, the chemical heterogeneity between BC and BS was revealed, establishing a comprehensive quality evaluation system encompassing microstructural features, spatial metabolite distribution, and elemental profiles. These findings offer a novel methodological framework for the authentication of Bupleurum species and the refinement of quality control standards.