The relative toxicity of the combined nonxylose components in prehydrolysates derived from three different lignocellulosic biomass feedstocks was determined. One woody (poplar) and two herbaceous (corn stover and switchgrass) feedstocks were dilute-acid pretreated under temperature and acid conditions chosen to optimize xylose recovery in the liquid prehydrolysate; xylose yields averaged 96, 89,and 87% of theoretical for switchgrass, corn stover, and poplar, respectively. Prehydrolysates from each feedstock were neutralized, adjusted to equivalent xylose concentrations, and bioassayed for toxicity, using a standardized fermentation protocol Mi with Pichia stipitis NRRL 11545. Full time-courses for ethanol production (30-60 h) clearly illustrate the distinct inhibitory effects of prehydrolysates from different feedstocks. The relative toxicity of the prehydrolysates, ranked in order of decreasing toxicity, is poplar-derived prehydrolysates > switchgrass-derived prehydrolysates > corn stover-derived prehydrolysates. The inhibition of ethanol production appeared to be the result of a general slowdown of yeast metabolism, rather than the result of the production of alternative, nonethanol end products. Ethanol yields averaged 74, 83, and 88% of control values for poplar, switchgrass, and corn stover prehydrolysates, respectively. Volumetric ethanol productivities (g ethanol L/h) averaged 32, 70, and 102% of control values for poplar, switchgrass, and corn stover prehydrolysates, respectively. Ethanol productivities correlated closely with acetate concentrations in the prehydrolysates; however, regression Lines correlating acetate concentrations and ethanol productivities were found to be feedstock-dependent.