AbstractAlthough many studies have been conducted on the wood property and chemical changes caused by thermal modification, little has been reported on the microstructural and topochemical changes occurring in the cell wall during heat treatment. In this study, poplar (Populus cathayaha) was treated within a temperature range from 180 to 220 °C for 4 h. Chemical analyses by Fourier transform infrared spectroscopy (FTIR) and nuclear magnetic resonance (NMR) indicated that heat treatment resulted in deacetylation of hemicelluloses and cleavage of lignin chains, thus generating new carbonyl and phenolic linkages. Transformation of matrix substances contributed to microstructural changes that appeared in clearly distorted and collapsed fiber and vessel walls along with the delamination of compound middle lamella (CML) and secondary walls (S), which showed a reduced capability to resist deformation. It was also observed by fluorescence microscopy (FM) and scanning electron microscope coupled with energy dispersive X-ray analysis (SEM-EDXA) that the concentration of lignin increased, probably because of the degradation of hemicelluloses and the generation of new carbonyl groups. These results on cell wall microstructure and topochemistry can help explain the altered wood properties revealed by dynamic mechanical analysis (DMA) and equilibrium moisture content (EMC) testing after heat treatment.