Conditioning of lignocellulosic hydrolysates with sulfur oxyanions, such as dithionite, is one of the most potent methods to improve the fermentability by counteracting effects of inhibitory by-products generated during hydrothermal pretreatment under acidic conditions. The effects of pH, treatment temperature, and dithionite dosage were explored in experiments with softwood hydrolysates, sodium dithionite, and Saccharomyces cerevisiae yeast. Treatments with dithionite at pH 5.5 or 8.5 gave similar results with regard to ethanol productivity and yield on initial glucose, and both were always at least ~20% higher than for treatment at pH 2.5. Experiments in the dithionite concentration range 5.0–12.5 mM and the temperature range 23–110◦ C indicated that treatment at around 75◦ C and using intermediate dithionite dosage was the best option (p ≤ 0.05). The investigation indicates that selection of the optimal temperature and dithionite dosage offers great benefits for the efficient fermentation of hydrolysates from lignin-rich biomass, such as softwood residues. © 2021 by the authors.