Two-stage sulfite steam pre-treatment achieves extraordinary bioethanol titer from simultaneously saccharification and fermentation of softwood whole slurries

Softwood biomass, which contains rich portion of readily fermentable sugars, is a type of abundant biomass in Canada and Nordic countries. However, its recalcitrant structure has severely limited its utilization in biocon-version. Sulfite treatment during steam pre-treatment to simultaneously enhance ease of hydrolysis and fermentation of softwood biomass has turned to be an attractive strategy to unlock the possibility of converting softwood to bioethanol. Our previous work showed applying one-stage sulfite treatment during the steam pre-treatment under neutral/alkaline conditions could maximize the sulfonation/delignification and hemicellulose sugar retention, while the use of acidic conditions could enhance the homogenous substrates size reduction. To take the advantages of both the neutral/alkaline and acidic conditions, we developed two-stage sulfite steam pre-treatment to maximize the sugar and ethanol production by utilizing Na2SO3 in the first alkaline steam pre-treatment stage and SO2 in a second acidic steam pre-treatment stage. The resulting substrates showed high sulfonation/delignification extent, nearly 100 % sugar recovery, homogeneous substrates size reduction and sufficient detoxification. Without extra hydrolysate detoxification and concentration, 160 g/L fermentable sugars were obtained from the hydrolysis of two-stage sulfite steam pretreated softwood whole slurry at 25% solid loading. And 80 g/L ethanol was generated by directly fermenting these sugars in the whole slurry hydrolysate without additional detoxification or concentration processes. This work significantly contributed to the devel-opment of the industrial relevant bioethanol production from softwood biomass.