SCALE-UP OF BIOETHANOL PRODUCTION FROM LIGNOCELLULOSIC BIOMASS HYDROLYSATE BY COFERMENTATION

Abstract

The present investigation aimed to explore the scale-up of bioethanol production from lignocellulosic biomass hydrolysate through co-fermentation. This study employed wild Zymomonas mobilis MTCC 91 and Scheffersomyces stipitis NCIM 3498 for the fermentation of hexose and pentose sugars, respectively. The objectives of this study were fourfold: optimizing the co-fermentation process, scaling up bioethanol production, achieving high cell density cultivation, and enhancing co-fermentation performance. Numerous wild-type microorganisms employed in ethanol production struggle to efficiently ferment pentose sugars, such as xylose and arabinose. Consequently, the adoption of diverse cultivation methods becomes essential, including co-culture, sequential cultures, and genetically engineered cultures. Sequential cultures offer several advantages, including stability, efficient xylose conversion, absence of compatibility issues between cultures, and catabolite repression. Hence, a simple and distinctive co-fermentation process was devised, employing sequential cultures. In the current sequential co-fermentation process, highly concentrated mixed sugars undergo conversion into bioethanol within a single bioreactor. This process involves the utilization of Zymomonas mobilis during stage 1 (glucose fermentation) and Scheffersomyces stipitis during stage 2 (xylose fermentation). The optimal mixed sugar concentration for fermentation was 160 g/L (comprising 100 g/L glucose and 60 g/L xylose). In the sequential co-fermentation process, during stage 1, an ethanol titer of 50.25 g/L was achieved with a yield of 0.50 g/g, and no residual glucose remained. However, in stage 2, there was minimal xylose conversion due to the presence of live Zymomonas mobilis cells. Upon removing these cells in stage 2, ethanol productivity resumed, resulting in an overall ethanol titer of 56.77 g/L with a yield of 0.36 g/g. Further improvements were observed when residual ethanol was removed along with the complete removal of Z. mobilis cells. On 80% removal of residual ethanol, overall ethanol titer and volumetric productivity were found to be 79.03 g/L and 1.09 g/L/h, respectively, and the same upon complete removal of residual ethanol was found to be 79.93 g/L and 1.82 g/L/h, respectively. Crucially, the fermentation performance achieved during the sequential co-fermentation of mixed sugars was more than 95% identical to that of separate glucose and xylose batch fermentation. Consequently, this outcome underscores the potential of the current sequential co-fermentation process for scale-up and large-scale bioethanol production from plant cell biomass. The present investigation aimed to explore the scale-up of bioethanol production from lignocellulosic biomass hydrolysate through co-fermentation. This study employed wild Zymomonas mobilis MTCC 91 and Scheffersomyces stipitis NCIM 3498 for the fermentation of hexose and pentose sugars, respectively. The objectives of this study were fourfold: optimizing the co-fermentation process, scaling up bioethanol production, achieving high cell density cultivation, and enhancing co-fermentation performance. Numerous wild-type microorganisms employed in ethanol production struggle to efficiently ferment pentose sugars, such as xylose and arabinose. Consequently, the adoption of diverse cultivation methods becomes essential, including co-culture, sequential cultures, and genetically engineered cultures. Sequential cultures offer several advantages, including stability, efficient xylose conversion, absence of compatibility issues between cultures, and catabolite repression. Hence, a simple and distinctive co-fermentation process was devised, employing sequential cultures. In the current sequential co-fermentation process, highly concentrated mixed sugars undergo conversion into bioethanol within a single bioreactor. This process involves the utilization of Zymomonas mobilis during stage 1 (glucose fermentation) and Scheffersomyces stipitis during stage 2 (xylose fermentation). The optimal mixed sugar concentration for fermentation was 160 g/L (comprising 100 g/L glucose and 60 g/L xylose). In the sequential co-fermentation process, during stage 1, an ethanol titer of 50.25 g/L was achieved with a yield of 0.50 g/g, and no residual glucose remained. However, in stage 2, there was minimal xylose conversion due to the presence of live Zymomonas mobilis cells. Upon removing these cells in stage 2, ethanol productivity resumed, resulting in an overall ethanol titer of 56.77 g/L with a yield of 0.36 g/g. Further improvements were observed when residual ethanol was removed along with the complete removal of Z. mobilis cells. On 80% removal of residual ethanol, overall ethanol titer and volumetric productivity were found to be 79.03 g/L and 1.09 g/L/h, respectively, and the same upon complete removal of residual ethanol was found to be 79.93 g/L and 1.82 g/L/h, respectively. Crucially, the fermentation performance achieved during the sequential co-fermentation of mixed sugars was more than 95% identical to that of separate glucose and xylose batch fermentation. Consequently, this outcome underscores the potential of the current sequential co-fermentation process for scale-up and large-scale bioethanol production from plant cell biomass.Notably, this fermentation strategy offers several advantages: it can significantly reduce byproduct inhibition, lower process costs, enhance productivity, and avoid the effects of carbon catabolite repression.