<p><i>Ferula ferulaeoides</i> is an endemic ephemeral medicinal plant in Xinjiang province of China. Visualizing the spatial distributions and dynamics of active metabolites is crucial for characterizing the metabolic networks of <i>F. ferulaeoides</i>, and this will also provide a highly informative approach to understand the complex molecular changes in the development of <i>F. ferulaeoides</i> resin duct (RD). In this study, matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was used to investigate the spatiotemporal changes of 3 terpenoids (<span>d</span>-limonene, myrcene, and tussilagone) and 4 coumarins (heraclenol, marmesin, 5,7-dihydroxycoumarin, and ammoresinol) in the 3 different development stages of RD, and further investigate the spatial distributions of 4 terpenoids (<span>d</span>-limonene, myrcene, tussilagone, and guaiol) and 6 metabolites in the coumarins synthesis pathway (umbelliferone, 7-demethylsuberosin, marmesin, heraclenol, 5,7-dihydroxycoumarin, and ammoresinol) within the stem of <i>F. ferulaeoides</i>. It was found that the signal intensities of 7 targeted metabolites were not observed in the lumen formation stage and gradually increased from the lumen enlargement stage to the mature stage of RD. Furthermore, the 2 monoterpenes presented the same spatial distribution, mainly located in the stem RD with high abundance. In contrast, guaiol (a sesquiterpene) exhibited a more uneven distribution in the stems, with lower signal intensity compared to the 2 monoterpenes. Meanwhile, the downstream metabolites involved in the biosynthetic pathway of coumarins, particularly 5,7-dihydroxycoumarin and ammoresinol, were specifically localized in the RD. In conclusion, these spatially-resolved metabolite data not only provide direct evidence that a series of metabolic alterations occurred in different development stages of <i>F. ferulaeoides</i> RD but also enhance our understanding of the <i>F. ferulaeoides</i> metabolic networks.</p> Graphical abstract <p></p>