Revisiting the Role of Nucleophiles in Suppressing Lignin Condensation and Enhancing Cellulose Saccharification during Organic Acid Pretreatment of Bamboo

Suppressing lignin condensation is crucial for the efficient deconstruction and utilization of lignocelluloses. However, the distinct stabilization behaviors of macromolecular lignin framework and low-molecular-weight lignin fragments remain unclear. How nucleophiles affect residual lignin–cellulase nonproductive adsorption has not been clearly elucidated. In this work, a cysteine-assisted formic acid (FA–Cys) pretreatment was used to separate lignin and improve the saccharification performance of bamboo biomass. Cysteine acted as a sulfur-based nucleophile to intercept reactive benzylic carbocations within the macromolecular lignin framework and aldehyde intermediates generated from β–O–4 cleavage of low-molecular-weight lignin fragments, thereby suppressing lignin condensation. The β–O–4 retention of recovered lignin increased from 25.9% to 89.6%, and the condensed structure decreased from 12.5 to nearly zero per 100 aromatic units. Cysteine decreased the maximum cellulase adsorption capacity of residual lignin from 10.5 to 4.5 mg/g, while excessive cysteine addition increased the adsorption capacity. High lignin removal (92.6%) and mitigated nonproductive adsorption of residual lignin resulted in an increase in glucose yield of FA–Cys-pretreated bamboo from 58.7% to 98.9%. This study advances the understanding of how nucleophiles suppress lignin condensation, providing a theoretical basis for the efficient separation of noncondensed lignin and enhanced cellulose saccharification of bamboo.