To compare the composition and performance of various lignocellulosic biomass hydrolysates as fermentation media, 8 hydrolysates were generated from a grass-like and a wood biomass. The hydrolysate preparation methods used were 1) dilute acid, 2) mild alkaline, 3) alkaline/peracetic acid, and 4) concentrated acid. These hydrolysates were fermented at 30°C, pH 5.0 using Saccharomyces cerevisiae CEN.PK113-7D as model strain. The growth in different hydrolysates varied in the aspects of lag-phase, growth rate, glucose consumption rate and ethanol production rate. Subsequently, 11 potential inhibitory compounds as described in literature were selected for further analysis. The concentrations of these compounds were determined in the time-samples of the 8 fermentations, using a novel analytical method, ethyl-chloroformate derivatization-GC-MS. Some of these compounds, e.g. furfural, decreased during the fermentation process, while others, such as formic and benzoic acid, remained almost constant. Inhibitory effect analysis of individual compound revealed that most of these compounds exhibit little effect at the concentrations detected in hydrolysates. Only furfural and benzoic acid clearly affected the growth of the model yeast: furfural elongated the lag-phase, while benzoic acid reduced the growth rate and biomass yield.