Abstract: Abstract: Heartwood formation involves the specific accumulation of substantial secondary metabolites within the inner wood tissues. Elucidating the distribution patterns of these metabolites across different xylem cell types (vessels, axial parenchyma cells, ray parenchyma cells and wood fibers) is crucial for deciphering the mechanisms underlying heartwood formation. In this study, the precious tree species Dalbergia odorifera was investigated using Ultra-Performance Liquid Chromatography-Mass Spectrometry (UPLC-Q-TOF-MS) to systematically analyze the distribution characteristics of four key secondary metabolites—Sativanone, Vestitone, (-)-Medicarpin, and Formononetin—across six radial zones of the xylem: Outer Sapwood (OSW), Middle Sapwood (MSW), Inner Sapwood (ISW), Outer Transition Zone (OTZ), Inner Transition Zone (ITZ), and Heartwood (HW). The results demonstrated a significant increasing trend in the content of all four secondary metabolites from the outer sapwood to the heartwood, indicating that heartwood formation is accompanied by continuous synthesis and accumulation of these compounds. Furthermore, Time-of-Flight Secondary Ion Mass Spectrometry (TOF-SIMS) was employed to resolve the spatial distribution patterns of flavonoid metabolites within xylem tissues. The analysis revealed that these characteristic metabolites are predominantly enriched in the heartwood region, with a non-uniform distribution pattern: signal intensity in heartwood fiber tissues was significantly stronger than that in parenchyma tissues.