Lignocelluloses are a diverse group of substrates  including cellulose, hemicellulose, lignin and extractives , which produce inhibitors during hydrolysis. The quantity of inhibitor depends not only on the origin of the material but also on the pre-treatment and hydrolysis method. Fermentation inhibitors, which inhibit yeast metabolism, include 5-hydroxymethyl furfural (HMF), 2-furaldehyde (furfural), phenolic compounds and weak acids for review, see [3–6]. Reportedly, weak acids exert a growth inhibitory effect by inflow of non-dissociated acid into the cytoplasm of the microorganism . HMF and furfural are consumed by S. cerevisiae, with consequent cost of ATP [8, 5]. The inhibitory effect of phenolic compounds on yeast metabolism remains under investigation. A feasible suggestion is that phenolic compounds degrade the cell membrane integrity, reducing the membrane’s efficacy as a selective barrier . To release high amounts of monosaccharides for fermentation, lignocelluloses (on account of their recalcitrant nature) often require harsh pre-treatment conditions. In most cases, the quantity of inhibitors increases with the severity of pre-treatment. Therefore, scientists seek mild pre-treatments that maximize the saccharification yields.
Besides containing growth inhibitors, lignocelluloses are generally scarce in nutrients and nutrient supplementation is thought to increase fermentation performance [10–12]. Earlier studies on such raw materials have identified lack of nitrogen as a crucial limiting factor in fermentations. Nitrogen is especially important for fermentative performance because it promotes cell proliferation [13, 14] and thereby ethanol yields, since the ethanol production rate is maximized in actively growing cells [15, 16].
Efficient distillation energy requires both high productivity and high ethanol titres. A prerequisite of ethanol production is high-gravity substrate, which theoretically yields at least 40–50 g L−1 ethanol . In practice, however, increasing the initial high dry matter content will also increase the concentrations of inhibitory compounds. The tolerance to inhibitors and the fermentation performance of cells depends on the nature of the substrate and the extent to which the fermenting organism is adapted to the specific challenges imposed by the substrate [18, 19]. Substrate stress affects the energy metabolism of the yeast cells, since many stress responsive processes depend on ATP availability . Some of the inhibitory compounds in lignocellulosic material appear to decrease the specific sugar uptake rate and the specific ethanol production rate [21, 5], both of which are highly correlated with ATP production. Adenine nucleotides also participate in numerous intracellular reactions, and their intracellular concentrations may greatly affect metabolism and yeast cell performance . The study of energy metabolism can therefore provide insights into the maintenance requirements of yeast cells in lignocellulosic fermentations.
To determine the re-usability of the yeast cells, we must know whether the cells can sustain high ethanol production rates under extended periods of stress and nutrient limitation. This is crucial for processes involving cell recycling or re-circulation. During cultivation, the ethanol production rate may not reflect the ethanol production capacity, because cell performance is reduced by the abovementioned substrate limitations. In this study, we measured the fermentation capacity of the cells after re-inoculation in a non-inhibitory nutritionally rich medium. Using this approach, we can detect whether the cells are irreversibly affected by the previous fermentation conditions.
To evaluate the different performance of yeast strains grown in different lignocellulosic substrates and the effect of nutrient addition, the fermentation performance, energy status and fermentative capacity of the strains were measured.