2,4-Bis(4-methoxyphenyl)-1,3,2,4-dithiadiphosphetane-2,4-disulfide (Lawesson's reagent, LR) reacted with pyrano[2,3-d]pyrimidine-2,4-(1H,3H)-dione derivatives in boiling acetonitrile to afford various pyrimido[5',4':5,6]pyrano[2,3-d][1,3,2]thiazaphosphinine derivatives. The structures of the target compounds were characterized by elemental analysis and spectral data. The biological activities of all synthesized compounds were tested against various microorganisms by the disk diffusion method. In general, the newly synthesized compounds showed good inhibitory effects against most of the applied microorganisms.