The present study focuses on the hydrothermal flow-through treatment of lignocellulosic biomass in a fixed-bed 3 L reactor. In combination with a sequential enzymatic hydrolysis, this process enables selective separation of the lignocellulose in its major constituents in minimal equipment. The aim of this work was to acquire detailed knowledge on the particle and bed structure, biomass degradation and non-ideal flow effects during the hydrothermal treatment. Correlations between the hemicellulose solubilization progress and changing fixed-bed characteristics were identified. This information is essential for the development of a realistic process model that considers hemicellulose reaction kinetics as well as mass and heat transfer separately. Hydrothermally pretreated wheat straw beds were analyzed using the tracer pulse response method at ambient conditions. Non-ideal flow was modeled by means of the dispersion model. Additionally, the effective porosity could be calculated from the residence time distribution curves. It was shown that the axial dispersion coefficient as well as the effective porosity were constant after 15 min of pretreatment. A linear relationship between the axial dispersion coefficient and the interstitial fluid velocity was found. These results allow simplifying the model by assuming constant values of the corresponding variables. The model can further be used to support the development of biorefinery plants at lab and production scale. (C) 2015 Elsevier B.V. All rights reserved.