Browsing by Author "Papoutsakis, E. Terry"

Now showing 1 - 4 of 4
  • Thumbnail Image
    Item
    Gas chromatography and gateway sensors for on-line state estimation of complex fermentations (butanol/acetone fermentations)
    (1984) McLaughlin, Joseph K.; Papoutsakis, E. Terry; Rudolph, Frederick B.; McIntire, Larry V.
    A bench-scale system for monitoring butanol/acetone fermentations was designed to demonstrate the use of on-line gas chromatography (GC) and the applicability of an "advanced" fermentation equation. Ultrafiltration was used to obtain sterilely a cell-free sample from a controlled fermentor for on-line GC analysis of ethanol, acetone, acetate, butanol, acetoin, and butyrate. The fermentor effluent gas was also analyzed by on-line GC for nitrogen, carbon monoxide, and carbon dioxide. Analytical data were automatically transmitted to a VAX 11/75 computer, for data archiving and fermentation equation calculations. The system was demonstrated in studies of pH and CO effects on the metabolism of Clostridium acetobutylicum. The on-line analytical systems provided excellent real time data. Also, three potentially useful gateway sensors were developed from the fermentation equation: glucose concentration, NADH2 from FdH2 and excess ATP. These demonstrated the usefulness of the fermentation equation for determination of intracellular and extracellular parameters in transient operation.
  • Thumbnail Image
    Item
    Investigation of the control of metabolic pathways in Clostridium acetobutylicum by the studies of glucose and non-glucose limitation, in vitro enzyme inhibition, and intermediary compound challenges in batch and continuous cultures
    (1984) Roos, Joseph William; Papoutsakis, E. Terry; Rudolph, Frederick B.; McIntire, Larry V.
    The fermentation of carbohydrates by Clostridium aoetobutylicum leads to the formation of acetate, acetoin, acetone, butanol, butyrate, ethanol, hydrogen, and carbon dioxide. The availability of ATP and NADH£ appears to be significant in determining the amount of products formed. Non-glucose limited, uncontrolled pH, batch cultures are capable of producing solvents. Increasing the ammonium to glucose ratio of these cultures resulted in an increase in acid formation and growth and a decrease in solvent yields. Maintaining a constant pH at 4.5 in batch cultures encouraged growth and glucose utilization resulting in glucose limited cultures. These ATP limited cultures gave high acid yields and small solvent yields. Butyraldehyde and acetoacetate influenced the metabolism of uncontrolled pH batch cultures. Their presence resulted in increased biomass formation but had mixed effects on acid and solvent production. A non-glucose limited continuous culture has been observed to produce butanol, acetone, butyrate and acetate, while under glucose limited conditions, continuous cultures produced primarily butyrate and acetate. In a glucose limited culture carbon monoxide was used to inhibit the hydrogenase activity. This inhibition resulted in an uptake of butyrate, a decrease in the rate of acetate production and a large increase in the specific rates of production of butanol, ethanol, and acetoin. Acetone was not affected.
  • Thumbnail Image
    Item
    Isolation, characterization and substrate-transport studies of a new, unique methylotroph
    (1984) Keuer, Thomas Alan; Papoutsakis, E. Terry; Storck, Roger L.; McIntire, Larry V.
    Methylotrophic bacteria which assimilate carbon via the Ribulose Monophosphate Pathway are bioenergetically superior to other methylotrophs. The dehydrogenases which catalyze the oxidation of formaldehyde to formate and formate to CO2 in RMP bacteria produce much of the ATP required for biosynthesis. A strain, designated T15, has been isolated on the basis of high In vitro activities of the above two key enzymes, and has been biochemically characterized. The new strain exhibits high yields (up to .63 g cells/g MeOH) and growth rates (up to .46 hr) in batch culture? however, the yields and growth rates in continuous culture are significantly lower. Study of the transport mechanisms has provided valuable insight into the relationship between substrate uptake and the growth characteristics of T15. Experiments with radio-labelled substrates have indicated that methanol enters the cells primarily by diffusion? consequently, the bacteria are not able to accumulate methanol internally in order to support efficient continuous growth. Formaldehyde, on the other hand, is accumulated by an active transport system which depends on the A pH component of the membrane proton-motive force. The formate uptake mechanism is also dependent on the ApH, but ‘is more complex, possibly due to the uncoupling effect of the organic acid on the cell membrane.
  • Thumbnail Image
    Item
    Shear stress effects on cultured hybridoma cells in a rotational couette viscometer
    (1989) Petersen, Johathan Franklin; Papoutsakis, E. Terry
    Cells growing in stirred bioreactors exist in a complex fluid mechanical environment. A number of forces act on the cells in the reactor, including fluid shear stresses. If agitation is sufficiently rapid, these forces may be lethal to the cells. In this study, the effects of well defined fluid shear on cell damage were investigated in a rotational couette viscometer. The shear sensitivity of the cells was modulated by the age of the culture. For cells that experience a prolonged stationary phase, the cells were quite sensitive to shear for both young and old cultures. If the stationary phase was short, the resistance to shear was higher throughout, and declined slightly with increasing culture age. The shear sensitivity of the cells was also modulated by specific components of the cytoskeleton. Disruption of the microfilaments made the cells more sensitive to shear, while disruption of the microtubules had no effect on shear sensitivity. Shear sensitivity also depended on energy metabolism in the cells. Inhibiting respiration increased shear sensitivity, and inhibiting glycolysis caused a further increase in shear sensitivity. Addition of fetal bovine serum to the culture medium made the cells more resistant to shear in a dose dependent manner. Addition of the pluronic polyol F68 to culture media had no effect. Polyethylene glycol increased shear sensitivity. Subjecting the cells to extreme agitation over an extended period resulted in a population of cells that grew more readily at high agitation rates. This subpopulation had a higher specific growth rate and less shear sensitivity than unselected cells.