Multi-omics and DESI-MSI reveal spatiotemporal chemical changes in soybean seeds during early germination

Soybean (Glycine max) germination is widely used to improve nutritional and functional quality, yet the earliest germination stage remains chemically under-characterized. Here, untargeted metabolomics, DDA-based proteomics, and desorption electrospray ionization mass spectrometry imaging (DESI-MSI) were integrated to investigate soybean seeds at 0, 1, and 2 d of germination. Early germination caused marked changes in metabolite and protein profiles, with differential metabolites mainly enriched in flavonoid, isoflavonoid, secondary-metabolite, and alpha-linolenic acid metabolism. Isoflavone-related compounds showed a clear day-1-centered response, whereas several lipid-derived metabolites remained elevated through day 2. Proteomics further indicated increased abundance of proteins related to translation, ribosome function, and central carbon metabolism. Integrated analysis revealed convergence between metabolite and protein changes at the KEGG pathway level and identified coordinated metabolite-protein modules. DESI-MSI added spatial evidence by showing tissue-associated localization of isoflavone-related signals. Overall, early germination was accompanied by coordinated temporal, molecular, and spatial changes in soybean seed chemistry rather than a simple bulk increase in functional metabolites, highlighting germination duration as a key variable for tuning soybean chemical quality.