RESUMEN
The total yield of ergosterol produced by the fermentation of the yeast Saccharomyces cerevisiae depends on the final amount of yeast biomass and the ergosterol content in the cells. At the same time ergosterol purity-defined as percentage of ergosterol in the total sterols in the yeast-is equally important for efficient downstream processing. This study investigated the development of both the ergosterol content and ergosterol purity in different physiological (metabolic) states of the microorganism S. cerevisiae with the aim of reaching maximal ergosterol productivity. To expose the yeast culture to different physiological states during fermentation an on-line inference of the current physiological state of the culture was used. The results achieved made it possible to design a new production strategy, which consists of two preferable metabolic states, oxidative-fermentative growth on glucose followed by oxidative growth on glucose and ethanol simultaneously. Experimental application of this strategy achieved a value of the total efficiency of ergosterol production (defined as product of ergosterol yield coefficient and volumetric productivity), 103.84 × 10-6 g L-1 h-1 , more than three times higher than with standard baker's yeast fed-batch cultivations, which attained in average 32.14 × 10-6 g L-1 h-1 . At the same time the final content of ergosterol in dry biomass was 2.43%, with a purity 86%. These results make the product obtained by the proposed control strategy suitable for effective down-stream processing. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:838-848, 2017.
Asunto(s)
Ergosterol/metabolismo , Saccharomyces cerevisiae/metabolismo , Biomasa , Reactores Biológicos/microbiología , Etanol/metabolismo , Fermentación/fisiologíaRESUMEN
The main objective of this work was to establish those factors either physical (power input) or chemical (limiting substrate or dilution rate) that enhance cell aggregation (biofilm or floc formation) and cell physiological state during aerobic continuous cultures of Bacillus licheniformis. Glucose-limited steady-state continuous cultures growing at a dilution rate between 0.64 and 0.87/h and 1,000 rpm (mean specific energy dissipation rate (epsilonT) = 6.5 W/kg), led to the formation of a thin biofilm on the vessel wall characterized by the presence of a high proportion of healthy cells in the broth (after aggregate disruption by sonication) defined as having intact polarized cytoplasmic membranes. An increased epsilonT (from 6.5 W/kg to 38 W/kg) was found to hinder cell aggregation under carbon limitation. The carbon recovery calculated from glucose indicated that additional extracellular polymer was being produced at dilution rates >0.87/h. B. licheniformis growth under nitrogen limitation led to floc formation which increased in size with dilution rate. Counter-intuitively the flocs became more substantial with an increase in epsilonT from 6.5 W/kg to 38 W/kg under nitrogen limitation. Indeed the best culture conditions for enhanced metabolically active cell aggregate formation was under nitrogen limitation at epsilonT = 6.5 W/kg (leading to floc formation), and under carbon limitation at a dilution rate of between 0.64 and 0.87/h, at epsilonT = 6.5 W/kg (leading to vessel wall biofilm formation). This information could be used to optimize culture conditions for improved cell aggregation and hence biomass separation, during thermophilic aerobic bioremediation processes.
Asunto(s)
Bacillus/citología , Bacillus/fisiología , Reactores Biológicos/microbiología , Técnicas de Cultivo de Célula/métodos , Glucosa/metabolismo , Modelos Biológicos , Nitrógeno/metabolismo , Aerobiosis/fisiología , Algoritmos , Biopelículas/crecimiento & desarrollo , Agregación Celular/fisiología , Proliferación Celular , Tamaño de la Célula , Supervivencia Celular , Simulación por Computador , Movimiento (Física)RESUMEN
Multi-parameter flow cytometry was used to monitor the population dynamics of Bacillus licheniformis continuous cultivations and the physiological responses to a starvation period and a glucose pulse. Using a mixture of two specific fluorescent stains, DiOC6(3) (3,3'-dihexylocarbocyanine iodide), and PI (propidium iodide), flow cytometric analysis revealed cell physiological heterogeneity. Four sub-populations of cells could be easily identified based on their differential fluorescent staining, these correspond to healthy cells (A) stained with DiOC6(3); cells or spores with a depolarised cytoplasmic membrane (B), no staining; cells with a permeabilised depolarised cytoplasmic membrane (C), stained with PI; and permeablised cells with a disrupted cytoplasmic membrane 'ghost cells' (D), stained with both DiOC6(3) and PI. Transmission electron micrographs of cells starved of energy showed different cell lysis process stages, highlighting 'ghost cells' which were associated with the double stained sub-population. It was shown, at the individual cell level, that there was a progressive inherent fluctuation in physiological heterogeneity in response to changing environmental conditions. All four sub-populations were shown to be present during glucose-limited continuous cultures, revealing a higher physiological stress level when compared with a glucose pulsed batch. A starvation period (batch without additional nutrients) increased the number of cells in certain sub-populations (cells with depolarised cytoplasmic membranes and cells with permeabilised depolarised cytoplasmic membranes), indicating that such stress may be caused by glucose limitation. Such information could be used to enhance process efficiency.
Asunto(s)
Algoritmos , Bacillus/citología , Bacillus/fisiología , Técnicas de Cultivo de Célula/métodos , Recuento de Colonia Microbiana/métodos , Citometría de Flujo/métodos , Glucosa/metabolismo , Interpretación de Imagen Asistida por Computador/métodos , Reactores Biológicos/microbiología , Proliferación Celular , CinéticaRESUMEN
A study has been made of thermophilic aerobic biodegradation of the liquid fraction of potato slops (distillation residue) from a rural distillery. The COD of this fraction ranged from 49 to 104 g O2/l, the main contributions to the COD coming from organic acids, reducing substances, and glycerol. It was found that biodegradation could be divided into the following stages: organic acids were removed first, followed by reducing substances and glycerol. The extent of removal varied according to the process temperature. At 50 degrees C, acetic and malic acids were removed completely, but the amount of isobutyric acid increased. At 60 degrees C, organic acid removal ranged from 51.2% (isobutyric acid) to 99.6% (lactic acid). Removals of glycerol and reducing substances were 86.2% and 87.4%, respectively. COD reduction was also temperature dependent, the highest removal efficiency (76.7%) being achieved at 60 degrees C. Dissolved oxygen may have limited the biodegradation process, as indicated by the DOT-versus-time profile.