Effect of controlled oxygen limitation on Candida shehatae physiology for ethanol production from xylose and glucose.

Carbon distribution and kinetics of Candida shehatae were studied in fed-batch fermentation with xylose or glucose (separately) as the carbon source in mineral medium. The fermentations were carried out in two phases, an aerobic phase dedicated to growth followed by an oxygen limitation phase dedicated to ethanol production. Oxygen limitation was quantified with an average specific oxygen uptake rate (OUR) varying between 0.30 and 2.48 mmolO(2) g dry cell weight (DCW)(-1) h(-1), the maximum value before the aerobic shift. The relations among respiration, growth, ethanol production and polyol production were investigated. It appeared that ethanol was produced to provide energy, and polyols (arabitol, ribitol, glycerol and xylitol) were produced to reoxidize NADH from assimilatory reactions and from the co-factor imbalance of the two-first enzymatic steps of xylose uptake. Hence, to manage carbon flux to ethanol production, oxygen limitation was a major controlled parameter; an oxygen limitation corresponding to an average specific OUR of 1.19 mmolO(2) g DCW(-1) h(-1) allowed maximization of the ethanol yield over xylose (0.327 g g(-1)), the average productivity (2.2 g l(-1) h(-1)) and the ethanol final titer (48.81 g l(-1)). For glucose fermentation, the ethanol yield over glucose was the highest (0.411 g g(-1)) when the specific OUR was low, corresponding to an average specific OUR of 0.30 mmolO(2) g DCW(-1) h(-1), whereas the average ethanol productivity and ethanol final titer reached the maximum values of 1.81 g l(-1) h(-1) and 54.19 g l(-1) when the specific OUR was the highest.

Dynamic microbial response under ethanol stress to monitor Saccharomyces cerevisiae activity in different initial physiological states.

Dynamic Saccharomyces cerevisiae responses to increasing ethanol stresses were investigated to monitor yeast viability and to optimize bioprocess performance when gradients occurred due to the specific configuration of multi-stage bioreactors with cell recycling or of large volume industrial bioreactors inducing chemical heterogeneities. Twelve fed-batch cultures were carried out with initial ethanol concentrations (P(in)) ranging from 5 g l(-1) to 110 g l(-1) with three different inoculums in different physiological states in terms of viability and quantity of ethanol produced (P(o)). For a given initial cell viability of 50%, the time to reach the maximum growth rate and maximum ethanol production rate was dependent on the difference P(in) - P(o). Whatever the initial physiological state, when the initial ethanol concentration P(in) reached 100 g l(-1), the yeasts died. Experimental results showed that the initial physiological state of the yeast was the major parameter to determine, the microorganisms' capacities to adapt and resist environmental changes.

Very high ethanol productivity in an innovative continuous two-stage bioreactor with cell recycle.

The performance of an innovative two-stage continuous bioreactor with cell recycle-potentially capable of giving very high ethanol productivity-was investigated. The first stage was dedicated to cell growth, whereas the second stage was dedicated to ethanol production. A high cell density was obtained by an ultrafiltration module coupled to the outlet of the second reactor. A recycle loop from the second stage to the first one was tested to improve cell viability and activity. Cultivations of Saccharomyces cerevisiae in mineral medium on glucose were performed at 30 degrees Celsius and pH 4. At steady state, total biomass concentrations of 59 and 157 gDCW l(-1) and ethanol concentrations of 31 and 65 g l(-1) were obtained in the first and second stage, respectively. The residual glucose concentration was 73 g l(-1) in the first stage and close to zero in the second stage. The present study shows that a very high ethanol productivity (up to 41 g l(-1) h(-1)) can indeed be obtained with complete conversion of the glucose and with a high ethanol titre (8.3 degrees GL) in the two-stage system.

Synergistic temperature and ethanol effect on Saccharomyces cerevisiae dynamic behaviour in ethanol bio-fuel production.

The impact of ethanol and temperature on the dynamic behaviour of Saccharomyces cerevisiae in ethanol biofuel production was studied using an isothermal fed-batch process at five different temperatures. Fermentation parameters and kinetics were quantified. The best performances were found at 30 and 33 degrees C around 120 g l(-1) ethanol produced in 30 h with a slight benefit for growth at 30 degrees C and for ethanol production at 33 degrees C. Glycerol formation, enhanced with increasing temperatures, was coupled with growth for all fermentations; whereas, a decoupling phenomenon occurred at 36 and 39 degrees C pointing out a possible role of glycerol in yeast thermal protection.

Aeration strategy: a need for very high ethanol performance in Saccharomyces cerevisiae fed-batch process.

In order to identify an optimal aeration strategy for intensifying bio-fuel ethanol production in fermentation processes where growth and production have to be managed simultaneously, we quantified the effect of aeration conditions--oxygen limited vs non limited culture (micro-aerobic vs aerobic culture)--on the dynamic behaviour of Saccharomyces cerevisiae cultivated in very high ethanol performance fed-batch cultures. Fermentation parameters and kinetics were established within a range of ethanol concentrations (up to 147 g l(-1)), which very few studies have addressed. Higher ethanol titres (147 vs 131 g l(-1) in 45 h) and average productivity (3.3 vs 2.6 g l(-1) h(-1)) were obtained in cultures without oxygen limitation. Compared to micro-aerobic culture, full aeration led to a 23% increase in the viable cell mass as a result of the concomitant increase in growth rate and yield, with lower ethanol inhibition. The second beneficial effect of aeration was better management of by-product production, with production of glycerol, the main by-product, being strongly reduced from 12 to 4 g l(-1). We demonstrate that aeration strategy is as much a determining factor as vitamin feeding (Alfenore et al. 2002) in very high ethanol performance (147 g l(-1) in 45 h) in order to achieve a highly competitive dynamic process.