Glandular Trichomes Serve as the Primary Storage and Functional Sites of Oridonin in Isodon rubescens Leaves: Spatial Localization, Quantitative Validation, and Bioactivity Correlation.

As a non-volatile diterpenoid, oridonin represents the major bioactive compound in the medicinal plant Isodon rubescens (Hemsl.) Hara. This compound exhibits a broad range of pharmacological activities, including potent anticancer effects against various tumor types, as well as antibacterial and anti-inflammatory properties. Although the pharmacological properties of oridonin have been extensively characterized, its exact tissue-level distribution in leaves has yet to be elucidated. In this study, histochemical staining, desorption electrospray ionization mass spectrometry imaging (DESI-MSI), and dichloromethane-targeted extraction coupled with high-performance liquid chromatography (HPLC) were employed to determine the cellular distribution of oridonin in I. rubescens leaves. Histochemical staining with Hydrochloric acid-vanillin (HCl-vanillin) revealed intense fluorescence signals exclusively in peltate glandular trichomes, with no detectable fluorescence in mesophyll cells. DESI-MSI analysis showed that the characteristic ion signal of oridonin (m/z = 387.15) exhibited a punctate distribution pattern closely matching the spatial arrangement of glandular trichomes. Quantitative HPLC analysis demonstrated that oridonin content in glandular trichome extracts accounted for 77.44% of that in whole-leaf extracts, whereas mesophyll extracts contained only 10.20%, suggesting that glandular trichomes serve as the primary storage site. Furthermore, bioactivity assays revealed that glandular trichome-enriched extracts exhibited significant antibacterial activity against Bacillus subtilis, Micrococcus luteus, and Staphylococcus aureus, and showed cytotoxic effects on A549 human lung adenocarcinoma cells, with activity levels positively correlated with oridonin content. These convergent lines of evidence provide evidence that glandular trichomes are the main accumulation and storage sites of oridonin in I. rubescens leaves, and that trichome-stored oridonin constitutes the primary material basis for the antibacterial and cytotoxic activities of this plant. This study provides a cellular-level basis for the quality evaluation and breeding of high-oridonin I. rubescens varieties.