(498a) Hydrolysate Fermentation Characterization for Xylose-Fermenting Saccharomyces Cerevisiae Strains Derived From Directed Evolution for Improved Xylose Utilization and Tolerance to Inhibitors

The goal of this work is to quantify the spectrum of pretreatment-derived compounds from alkaline hydrogen peroxide (AHP) pretreatment of grasses, to quantify their impact on fermentation of high sugar content hydrolysates on Saccharomyces cerevisiae strains engineered to ferment xylose by introduction of xylose fermentation capabilities by introduction of either xylose reductase and xylitol dehydrogenase or the xylose isomerase followed by evolution for improved xylose utilization and AHP inhibitor tolerance, and to demonstrate improved performance and successful co-fermentation of undetoxified hydrolysates.  Specifically, we were able to determine that the classes of inhibitors included inorganics (Na⁺ from NaOH and sulfate used for pH adjustment) and solubilized organics (aromatics, acetate, and oxidation prodcucts including formate).  It was found that inorganics were not significantly inhibitory at concentrations of 0.5 M Na⁺ which would be present in hydrolysates from high-solids pretreatment and hydrolysis.  We were able to identify the soluble aromatics and especially the hydroxycinnamic acids as the major contributor to fermentation inhibition.  We are able to demonstrated complete sugar utilization from undetoxified AHP corn stover hydrolysates from high-solids pretreatment and enzymatic hydrolysis containing more than 120 g/L of glucose and xylose with conversion to ethanol at high yields with minimal byproduct formation.