Statistical Optimization of a Trichoderma-based Enzyme Combination for Saccharification of Steam-exploded Lignocellulosic Biomass

Abstract

This study aimed to determine the optimal enzyme combination conditions for improving the saccharification efficiency of softwood biomass (Larix kaempferi). For this purpose, cellulase derived from Trichoderma sp. KMF006 was combined with a commercial enzyme (Cellic® CTec3). Comparative hydrolysis experiments with individual enzymes showed that L. kaempferi exhibited a lower glucose yield than hardwood, suggesting the need for a complementary enzyme combination. A Plackett-Burman Design (PBD) was used to identify significant variables, including substrate concentration, enzyme loading, pH, and the KMF006 blending ratio. The significant factors were further optimized using a Box-Behnken Design (BBD). The optimal conditions were determined to be a substrate concentration of 9% (w/v), enzyme loading of 60 FPU/g-glucan, pH of 6.0, and the KMF006 blending ratio of 25.5%. The predicted maximum glucose yield under these conditions was 63.9%, representing a 21.8% increase compared to CTec3 alone and a 32.4% increase compared to KMF006 alone. These results suggest that up to 25% of the commercial enzyme dosage can be substituted with KMF006 without compromising hydrolysis performance. Overall, this study demonstrates the feasibility of an enzyme combination approach for enhancing softwood saccharification.