Significant improvements in pretreatments and a deeper understanding of changes in the physical structure and chemical composition of pretreated lignocellulosic materials are required to guarantee the technical and economic sustainability of second-generation ethanol technologies. In present work, several biophysical techniques were applied to characterize sugarcane bagasse samples that have been subjected to autocatalytic, phosphoric acid and sulfuric acid catalyzed steam explosion under equivalent combined severity factors (CSF). Confocal laser scanning microscopy and fluorescence lifetime imaging studies demonstrated unequal changes in lignin distribution in the plant cell wall that could be nicely correlated with substrate susceptibility to enzymatic hydrolysis. Furthermore, solid state nuclear magnetic resonance corroborated these observations by showing the differences in biomass composition primarily associated with hemicellulose removal, whereas X-ray diffraction analysis revealed changes in crystallinity indexes and average crystallite sizes. Thus, the impact of pretreatment temperature on the physical structure and chemical composition of CSF-equivalent steam-exploded sugarcane bagasse was identified as the key factor for developing substrate accessibility. This calls for caution in using combined severity factor as a chief parameter for comparison of different pretreatment conditions.
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