Box-Behnken design for the optimization of bioethanol production from rice straw and sugarcane bagasse by newly isolated Pichia occidentalis strain AS.2

This study investigated bioethanol production from rice straw (RS) and sugarcane bagasse (SCB) which containing 72.8 and 73.2% holocellulose, 56.8 and 58.6% α-cellulose, and 14.9 and 25.1% lignin for RS and SCB, respectively. To eliminate the lignin content, different pretreatment conditions, such as hot water, dilute acid, and acid-alkali, were designed. Acid-alkali was characterized as the best pretreatment for removing ∼79 and 70% of lignin, α-cellulose increased 91.4 and 91%, and holocellulose reached 90.8 and 90% for RS and SCB, respectively. The results revealed that acid-alkali was highly efficient than other pretreatment used for both RS and SCB. After enzymatic hydrolysis of acid-alkali-treated RS and SCB with cellulase, glucose concentrations reached 45 and 42 g/l, respectively. Pichia occidentalis AS.2 was isolated and identified based on 18S rRNA sequencing as a bioethanol producer. Maximization of bioethanol production by P. occidentalis AS.2 using the resulting glucose as a carbon source from RS and SCB was studied using an experimental design. The pH, incubation period, and inoculum size were optimized using Box-Behnken designs (BBD), the final conditions for bioethanol production used 100 g/l acid-alkali-treated fibers, 10 ml cellulase enzyme at 50°C for 5 days at 75 rpm for enzymatic hydrolysis. After time consumed and adjusting the pH to 6, the mixture was inoculated with 2.5% P. occidentalis AS.2 and incubated at 35°C for 24 h at 200 rpm to increase the bioethanol yield by 1.39-fold to 23.7 and 21.4 g/l compared to initial production (17 and 15.3 g/l) between RS and SCB, respectively.