Allelopathic inhibitors in seeds kernel cause dormancy variation between hard and soft Sapindus mukorossi seeds

Seed dormancy helps seeds to survive in nature but often reduces uniform germination under cultivation. In Sapindus mukorossi , seed dormancy is influenced by both physical and chemical factors. Hard seeds of S. mukorossi , which remain firm even after prolonged soaking, germinate earlier, while soft seeds that soften after soaking exhibit delayed germination. To unravel the mechanisms underlying dormancy variation, we combined allelopathic bioassays, UPLC–MS/MS–based metabolite profiling, and SEM-guided anatomical characterization of the seed coat. Bioassay experiments revealed that soft-seed kernel extracts of S. mukorossi exhibited strong allelopathic inhibitory effect on Chinese cabbage seeds germination. The 100% soft‑seed kernel extract (100 g/L) reduced cabbage seeds germination up to 6%, whereas the 100% hard-seed kernel extract showed much weaker allelopathic effects, with germination reaching up to 100%. Root growth was completely inhibited by the 100% soft-seed kernel extract, while soft-seed coat extracts permitted limited root elongation (up to 2.9 cm). Metabolomic profiling identified 1,120 metabolites in soft and hard seed kernels, of which 129 differed significantly between the two tissue types. Soft seeds kernels were notably enriched in 33 unique flavonoids and 11 unique alkaloids, including Hesperetin-7-O-glucoside, (-)-epicatechin, phlorizin derivatives, strictamine, methyl dioxindole-3-acetate, hexadecanamide, and caffeine. These compounds are candidate allelochemicals that may contribute to the inhibition of cabbage seed germination and seedling growth observed with the crude extracts. SEM showed that soft seeds had a rough outer surface with palisade layers containing spaces, crystal-like formations, and a porous endotesta. In contrast, hard seeds exhibited a porous outer surface but a highly compact palisade layer and interconnected endotesta. Our results suggest that delayed germination in soft S. mukorossi seeds is primarily associated with allelopathic inhibitors accumulated in the seed kernel rather than by physical restrictions of the seed coat. These findings highlight the importance of chemical factors in regulating dormancy in this species.