Combining treatments to improve the fermentation of sugarcane bagasse hydrolysates by ethanologenic Escherichia coli LY180

• High pH treatment (NH4OH) is the best single treatment to improve hydrolysate fermentation.
• Combined treatments were complimentary, incrementally improving fermentation.
• Vacuum removal of furans, high pH, laccase treatment, and bisulfite addition eliminated hydrolysate toxicity.
• After these 4 treatments, fermentations with treated hydrolysate were equivalent to laboratory sugars.

Inhibitory side products from dilute acid pretreatment is a major challenge for conversion of lignocellulose into ethanol. Six strategies to detoxify sugarcane hydrolysates were investigated alone, and in combinations (vacuum evaporation of volatiles, high pH treatment with ammonia, laccase, bisulfite, microaeration, and inoculum size). High pH was the most beneficial single treatment, increasing the minimum inhibitory concentration (measured by ethanol production) from 15% (control) to 70% hydrolysate. Combining treatments provided incremental improvements, consistent with different modes of action and multiple inhibitory compounds. Screening toxicity using tube cultures proved to be an excellent predictor of relative performance in pH-controlled fermenters. A combination of treatments (vacuum evaporation, laccase, high pH, bisulfite, microaeration) completely eliminated all inhibitory activity present in hydrolysate. With this combination, fermentation of hemicellulose sugars (90% hydrolysate) to ethanol was complete within 48 h, identical to the fermentation of laboratory xylose (50 g/L) in AM1 mineral salts medium (without hydrolysate).