Development of a Computed Tomography (CT) Scanning Approach to Quantify Biofilm Thickness in Biofilm Reactors

Quantitative assessment of biofilm thickness in biofilm reactors is essential for monitoring, understanding, and optimising the bioconversion processes. However, conventional characterization methods are often destructive and unsuitable for intact reactor systems. This study presents a non-destructive, computed tomography (CT)-based workflow for quantification of biofilm thickness, volume, and surface-area distribution in an operating Trickle-Bed Reactor (TBR). A lab-scale syngas-fed TBR (220 mL) containing polymeric packing with a mature biofilm was investigated in situ using a Nikon XT H 225 micro-CT system. Image segmentation based on thresholding allowed to differentiate biofilm, packing material, and reactor wall due to different X-ray attenuation. Quantitative analysis revealed biofilm thicknesses of 1.1–1.3 mm and volumetric occupancies of 9–13%, increasing towards the reactor bottom—consistent with expected hydrodynamic gradients. The CT procedure preserved reactor integrity and microbial activity, confirming its non-invasive nature and suitability for repeated imaging. The developed workflow provides a reliable and adaptable approach for three-dimensional biofilm characterization in intact reactors and can be extended to other biofilm-based systems. Integration of the developed CT scanning method with process monitoring and modelling will further advance understanding and optimisation of biofilm-based processes. • Micro-CT workflow quantifies biofilm thickness in biofilm reactors • Non-destructive, in situ method preserves reactor and biofilm structure • 3D imaging separates biofilm, packing, and wall without contrast agents • Image analysis enables quantitative assessment of biofilm spatial heterogeneity