Effects and management of lactobacilli in yeast-catalyzed ethanol fermentations

Abstract

This thesis focuses on the effects of lactobacilli and their end-products, lactic acid and acetic acid, on 'Saccharomyces cerevisiae' growth and fermentation, and on antimicrobials used to manage such contaminants. To assess the effects of the bacteria, normal gravity (22-24 g/100 ml dissolved solids) wheat mashes inoculated with yeast at ~106 colony forming units (CFU)/ml were deliberately infected (coinoculated) with each of five industrially important strains of lactobacilli at ~10 5, ~106, ~107, ~10 8, and ~109 CFU/ml. Controls with yeast alone or with bacteria alone (~107 CFU/ml) were included. End-products, yeast growth and fermentation rates were monitored. Results indicated that production of lactic acid by lactobacilli and suspected competition of the bacteria with yeast cells for essential growth factors in the fermenting medium were the major reasons for reductions in yeast growth and decreases in final ethanol yield. A chemically defined minimal medium was used to determine the effects of added acetic and lactic acid, and their mode of action on two strains of ' S. cerevisiae'. The effects of these two acids on yeast intracellular pH (pHi), plasma membrane H+-ATPase activity and on the plasma membrane lipid composition were studied. It was found that the specific growth rates ([mu]) of the two yeast strains decreased exponentially (R2 > 0.9) as the concentrations of acetic or lactic acid were increased. Acetic and lactic acids synergistically reduced the specific growth rate of yeast. Acetic acid caused the yeast cell to expend ATP to pump out excess protons that result from the passive diffusion of the acid into the cell at medium pH (pHe) followed by its dissociation within the cell as a result of higher pHi. Lactic acid (0.5 % w/v) caused intracellular acidification (which could lead to arrest in glycolytic flux) as a result of a significant decrease (P = 0.05) in the plasma membrane H +-ATPase activity. Moreover, the plasma membrane fluidity was reduced due to decrease in unsaturated fatty acyl residues. Among the antimicrobials studied, urea hydrogen peroxide (UHP) was superior compared to stabilized chlorine dioxide and nisin, but its bactericidal activity was greatly affected by the presence of particulate matter. When used near 30 mmoles/L (in unclarified mash), in addition to its bactericidal effect, UHP provided near optimum levels of assimilable nitrogen and oxygen that aided in vigorous yeast fermentation. This process was patented.