The Bo Phloi gem field in Kanchanaburi Province, Western Thailand, is closely associated with Cenozoic basalts. Blue and yellow sapphire, black spinel, and minor zircon have been mined for over three decades. The mineral inclusions observed in sapphire samples are alkali feldspar, nepheline, hercynitic spinel, zircon, manganiferous ilmenite, silica-rich enstatite, almandine–pyrope garnet, monazite, calcite, sapphirine, biotite–phlogopite mica, and staurolite. Based on their geochemical affinity, these mineral inclusions can be categorized into two main groups: felsic alkaline and contact-metamorphic, which appear to have originated from different processes. These inclusions provide new evidence for proposing a bimodal genetic model. Felsic alkaline origin is evidenced by the occurrence of a felsic alkaline inclusion suite and the REE geochemistry of sapphire-associated zircon, which indicates that most of the sapphires crystallized from a high-alkali felsic melt (probably, in the lower crust). Contact-metamorphic origin is evidenced by the presence of a contact-metamorphic inclusion suite, suggesting that some of these sapphires might also have originated from metasomatized crustal rocks and a contaminated melt along the contact zone of a basaltic intrusion (probably, in the upper mantle or lower crust).