Lignocellulosic biomass is one of the most abundant raw materials available on earth, and the study of lignocellulose components is required for the production of second-generation biofuels. Fourier transform infrared spectroscopy (FTIR) has a demonstrated potential as a costeffective and efficient method to distinguish between lignocellulose specimens. This study compared three FTIR modes-attenuated total reflectance (ATR), diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS), and Transmission-FTIR-in their ability to distinguish samples of different lignocellulose species at varying grain sizes, as well as before and after enzyme treatment. The reproducibility among replicates and the separation between different sample groups was assessed using an adjusted "separation/scatter" metric calculated from the scores of principal component analysis (PCA). Attenuated total reflectance was most frequently the best method due to its least amount of variance among sample replicates. However, Transmission-FTIR was better than ATR for certain particle sizes or enzyme treatments. Diffuse reflectance infrared Fourier transform spectroscopy was repeatedly inferior to ATR and Transmission-FTIR, especially in terms of variability. This work provided insight into the best mode of FTIR for characterizing lignocellulose powders. Future work should test the robustness of these results with a wider range of wood species, particle sizes, enzymes concentrations, and reaction conditions.

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