Tailoring a cellulolytic enzyme cocktail for efficient hydrolysis of mildly pretreated lignocellulosic biomass

Commercially available cellulase cocktails frequently demonstrate high efficiency in hydrolyzing easily digestible pretreated biomass, which often lacks hemicellulose and/or lignin fractions. However, the challenge arises with enzymatic hydrolysis of mildly pretreated lignocellulosic biomasses, which contain cellulose, hemicellulose and lignin in high proportions. This study aimed to address this question by evaluating the supplementation of a commercial cellulolytic cocktail with accessory hemicellulases and two additives (H₂O₂ and Tween® 80). Statistical optimization methods were employed to enhance the release of glucose and xylose from mildly pretreated sugarcane bagasse. The optimized supplement composition resulted in the production of 304 and 124 mg g^-1 DM of glucose and xylose, respectively, significantly increasing glucose release by 84% and xylose release by 94% compared to using only the cellulolytic cocktail. This enhancement might be attributed to a coordinated hemicellulases action degrading hemicellulose, creating more space for cellulase activity, potentially boosted by the presence of H₂O₂ and Tween® 80. However, the addition of different concentrations of H₂O₂ in combination with hemicellulase and Tween® 80 did not result a significant difference on sugar release, which could be attributed to the limited range of concentrations studied (5 to 65 µM). The results obtained in this study using the mix of three supplements were also compared to the addition of only hemicellulase and only Tween® 80 to the cellulolytic cocktail. A significant increase in glucose release of 39% and 41%, respectively, was observed when using the optimized combination. For xylose, the increase was 38% and 41%, respectively. This study underscores the substantial potential in optimizing enzyme cocktails for the hydrolysis of mildly pretreated lignocellulosic biomass by using enzymes and additive combinations tailored to the specific biomass composition.