Real Time and Quantitative Imaging of Lignocellulosic Films Hydrolysis by Atomic Force Microscopy Reveals Lignin Recalcitrance at Nanoscale

Lignocellulosic biomass is considered as a sustainable source of energy and chemicals, but its recalcitrance to bioconversion still limits its use. In this paper, a strategy based on two aspects was developed to improve our knowledge on the lignin recalcitrance to enzymatic hydrolysis. First, lignocellulosic films of cellulose nanofibrils (CNFs) with increasing content of lignin (up to 40%) were prepared. Thanks to in situ real time Atomic Force Microscopy (AFM) measurements during the hydrolysis and by comparison with biochemical assays, the use of such films allows to fully assess the importance of the lignin content and of the arrangement between CNFs and lignin on the hydrolysis efficiency. In a second time, contrary to other studies by AFM which only followed a specific structure during enzymatic processes mostly on simple systems (CNFs or cellulose nanocrystals), a quantitative analysis of in-situ time-lapse measurements was developed. It enables to accurately address lignocellulosic biomass recalci...