Sequential ultrasonic and hydrothermal pretreatment enhances cell wall degradation and enzymatic hydrolysis of pine and yellow poplar

Efficient and environmentally friendly pretreatment is essential to overcome the recalcitrance of lignocellulosic biomass and improve enzymatic hydrolysis. In this study, ultrasonic and hydrothermal pretreatments were applied individually and sequentially to pine and yellow poplar to evaluate their effects on cell wall degradation and enzymatic hydrolysis. For both pine and yellow poplar, the highest hemicellulose degradation occurred when ultrasonic and 190°C hydrothermal pretreatments were applied sequentially. The glucan content was 55.77 % for pine and 57.60 % for yellow poplar, with cellulose to glucose conversion rates of 41.62 % and 89.48 %, respectively. Hemicellulose degradation at the cell wall level was observed using transmission electron microscopy-immunogold labeling method, revealing a decrease in xylan epitopes in yellow poplar and mannan epitopes in pine during the sequential pretreatments. Confocal laser scanning microscopy observations revealed that lignin autofluorescence intensity varied depending on the pretreatment method, reflecting differences in structural and compositional modifications of lignin. Field-emission scanning electron microscopy analysis provided detailed insights into the morphological alterations of biomass after pretreatment and effectively visualized the disruption and degradation of cellulose microfibril orientation at the cell wall level following enzymatic hydrolysis. This integrated approach highlights the merit of sequential ultrasonic and hydrothermal pretreatment as an effective and green strategy for enhancing cell wall degradation and enzymatic hydrolysis efficiency in lignocellulosic biomass. • Sequential ultrasonic-hydrothermal pretreatment enhanced enzymatic hydrolysis efficiency. • Hemicellulose and lignin degradation at the cell wall level was confirmed by immuno-TEM, FE-SEM, and CLSM. • Immunolabeling showed reduced xylan and mannan epitope densities after pretreatment. • Lignin autofluorescence intensity increased following sequential pretreatment.