Combined substrate, enzyme and yeast feed in simultaneous saccharification and fermentation allow bioethanol production from pretreated spruce biomass at high solids loadings

Background: Economically feasible cellulosic ethanol production requires that the process can be operated at high solid loadings, which currently imparts technical challenges including inefficient mixing leading to heat and mass transfer limitations and high concentrations of inhibitory compounds hindering microbial activity during simultaneous saccharification and fermentation (SSF) process. Consequently, there is a need to develop cost effective processes overcoming the challenges when working at high solid loadings. Results: In this study we have modified the yeast cultivation procedure and designed a SSF process to address some of the challenges at high water insoluble solids (WIS) content. The slurry of non-detoxified pretreated spruce when used in a batch SSF at 19% (w/w) WIS was found to be inhibitory to Saccharomyces cerevisiae Thermosacc that produced 2 g l(-1) of ethanol. In order to reduce the inhibitory effect, the non-washed solid fraction containing reduced amount of inhibitors compared to the slurry was used in the SSF. Further, the cells were cultivated in the liquid fraction of pretreated spruce in a continuous culture wherein the outflow of cell suspension was used as cell feed to the SSF reactor in order to maintain the metabolic state of the cell. Enhanced cell viability was observed with cell, enzyme and substrate feed in a SSF producing 40 g l(-1) ethanol after 96 h corresponding to 53% of theoretical yield based on available hexose sugars compared to 28 g l(-1) ethanol in SSF with enzyme and substrate feed but no cell feed resulting in 37% of theoretical yield at a high solids loading of 20% (w/w) WIS content. The fed-batch SSF also significantly eased the mixing, which is usually challenging in batch SSF at high solids loading. Conclusions: A simple modification of the cell cultivation procedure together with a combination of yeast, enzyme and substrate feed in a fed-batch SSF process, made it possible to operate at high solids loadings in a conventional bioreactor. The proposed process strategy significantly increased the yeast cell viability and overall ethanol yield. It was also possible to obtain 4% (w/v) ethanol concentration, which is a minimum requirement for an economical distillation process.