Recent studies have highlighted the diverse roles of extracellular vesicles, which have been detected in natural products such as ginger and ginseng. This study examined whether traditional Japanese Kampo medicines also contain extracellular vesicles. The drug substance (i.e., spray-dried powder from hot-water extracts of crude drugs) of the Kampo medicine Ninjin’yoeito was suspended in distilled water and fractionated by size exclusion chromatography to isolate nano-sized particles. Nanoparticle tracking analysis and electron microscopy confirmed the particle size distribution and morphology of the product, confirming the presence of exosome-like nanoparticles in the Kampo medicine preparation. Nanoparticle measurements and electron microscopy revealed vesicle-like structures consistent with the known characteristics of exosomes, and proteomic analysis supported their biochemical identity. Lectin array profiling revealed the abundance of high-mannose N-glycans on their surface. These nanoparticles were internalized by cells, and miRNA analysis indicated the presence of multiple miRNA types within the nanoparticles. This is the first report demonstrating the presence and characteristics of exosome-like nanoparticles in Kampo medicines. Although most active components of Kampo medicines have traditionally been considered small molecules, our findings suggest that extracellular vesicles can also serve as potential bioactive components. • Exosome-like nanoparticles were detected in the drug substance of Kampo medicine Ninjin’yoeito • Integrated morphological, proteomic, and miRNA‑based analyses confirmed the identity of the exosome‑like nanoparticles • Lectin array demonstrated a predominance of high-mannose N-glycans on the nanoparticle surface • Fluorescence microscopy confirmed the uptake of the nanoparticles by mouse macrophage-like cells • Multiple miRNAs associated with extracellular vesicles were identified within the nanoparticles
Tsuchiya et al. (Wed,) studied this question.