Deep Eutectic Solvent Pretreatment of Transgenic Biomass With Increased C₆C₁ Lignin Monomers

The complex and heterogeneous polyphenolic structure of lignin confers recalcitrance to plant cell walls and challenges biomass processing for agroindustrial applications. Recently, significant efforts have been made to alter lignin composition to overcome its inherent intractability. In this work, to overcome technical difficulties related to biomass recalcitrance, we report an integrated strategy combining biomass genetic engineering with a pretreatment using a bio-derived deep eutectic solvent (DES). In particular, we employed biomass from an Arabidopsis line that expressed a bacterial hydroxycinnamoyl-CoA hydratase-lyase (HCHL) in lignifying tissues, which results in the accumulation of unusual C₆C₁ lignin monomers and a slight decrease in lignin molecular weight. The transgenic biomass was pretreated with renewable DES that can be synthesized from lignin-derived phenols. Biomass from the HCHL plant line containing C₆C₁ monomers showed increased pretreatment efficiency and released more fermentable sugars up to 34% compared to WT biomass. The enhanced biomass saccharification of the HCHL line is likely due to a reduction of lignin recalcitrance caused by the overproduction of C₆C₁ aromatics that act as degree of polymerization (DP) reducers and higher chemical reactivity of lignin structures with such C₆C₁ aromatics. Overall, our findings demonstrate that strategic plant genetic engineering, along with renewable DES pretreatment, could enable the development of sustainable biorefinery.