Evaluation of Saccharomyces cerevisiae modified via CRISPR/Cas9 as a cellulosic platform microorganism in simultaneously saccharification and fermentation processes.

The nonrenewable character and deleterious effects of fossil fuels foster the need for cleaner and more inexhaustible energy sources, such as bioethanol. Especially from lignocellulosic biomasses. However, the economic viability of this product in the market depends on process optimization and cost reduction. This research applied a sequential experimental project to investigate the process of enzymatic saccharification and simultaneous fermentation to produce ethanol with sugarcane bagasse. The differential of the work was the application of the strain of Saccharomyces cerevisiae AGY001 which was improved by evolutionary engineering to become thermotolerant and by a heterologous expression based on genomic integration by CRISPR/Cas9 to produce endoglucanase and ß-glucosidase (AsENDO-AsBGL). The maximum ethanol yield found was 89% of the maximum theoretical yield (released sugars), obtained at temperature concentrations, sugarcane bagasse and inoculum at 40 °C, 16.5%, and 4.0 g/L, respectively (12.5 FPU/g bagasse). The mathematical model obtained can predict approximately 83% of the data set with 95% confidence. Therefore, these findings demonstrated the potential of sugarcane bagasse and S. cerevisiae AGY001 strain (CRISPR/Cas9 modified) in bioethanol production without the need for impractical selection media on an industrial scale, in addition to providing useful insights for the development of SSF processes.

Enzymatic hydrolysis of sugarcane bagasse pretreated with acid or alkali

The aim of this study was to evaluate the performance of enzymatic hydrolysis of acid or alkali pretreated sugarcane bagasse for the production of fermentable sugars. The first step consisted of selection of commercial enzymes presenting the highest cellulolytic activities. After selection of four enzymes: HPL, CL, P1 and P4, their performances were tested in the bagasse pretreated with acid and alkali. The sugar content of the hydrolysates was analyzed by anion exchange liquid chromatography. Data showed that the joint action of 0.5 percent acid pretreatment, 121ºC, 30 minutes and enzyme CL provides the best results, 67.25 g of hexose and 148.13g of pentose per kg of dry bagasse.