Effects of microorganisms on effective oxygen diffusion coefficients and solubilities in fermentation media

103

The growth of microorganisms may be limited by operating conditions which provide an inadequate supply of oxygen. To determine the oxygen-transfer capacities of small-scale bioreactors such as shaking flasks, test tubes, and microtiter plates, a noninvasive easy-to-use optical method based on sulfite oxidation has been developed. The model system of sodium sulfite was first optimized in shaking-flask experiments for this special application. The reaction conditions (pH, buffer, and catalyst concentration) were adjusted to obtain a constant oxygen transfer rate for the whole period of the sulfite oxidation reaction. The sharp decrease of the pH at the end of the oxidation, which is typical for this reaction, is visualized by adding a pH dye and used to measure the length of the reaction period. The oxygen-transfer capacity can then be calculated by the oxygen consumed during the complete stoichiometric transformation of sodium sulfite and the visually determined reaction time. The suitability of this optical measuring method for the determination of oxygen-transfer capacities in small-scale bioreactors was confirmed with an independent physical method applying an oxygen electrode. The correlation factor for the maximum oxygen-transfer capacity between the chemical model system and a culture of Pseudomonas putida CA-3 was determined in shaking flasks. The newly developed optical measuring method was finally used for the determination of oxygen-transfer capacities of different types of transparent small-scale bioreactors.

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 97%

Optical method for the determination of the oxygen‐transfer capacity of small bioreactors based on sulfite oxidation

Hermann

Walther

Maier

et al. 2001

103

“…The composition of the liquid medium, i.e., the concentration of dissolved compounds, also influences the gasliquid mass transfer characteristics. Higher concentrations or ionic strength lead to a lower oxygen solubility (L O2 ) (Schumpe, 1993;Weisenberger and Schumpe, 1996) and lower mass transfer coefficients (k L ) (Linek et al, 1970;Zieminski and Whittemore, 1971) due to smaller diffusion coefficients (D O2 ) (Akita, 1981;Ho and Ju, 1988). Up to now, very little is known about the effects of liquid viscosity on the oxygen transfer rate.…”

Section: Sulfite Oxidationmentioning

confidence: 95%

Characterization of gas–liquid mass transfer phenomena in microtiter plates

Hermann

Lehmann

Büchs

2002

195

215

Gas-liquid mass transfer properties of shaken 96-well microtiter plates were characterized using a recently described method. The maximum oxygen transfer capacity (OTR(max)), the specific mass transfer area (a), and the mass transfer coefficient (k(L)) in a single well were determined at different shaking intensities (different shaking frequencies and shaking diameters at constant filling volume) and different filling volumes by means of sulfite oxidation as a chemical model system. The shape (round and square cross-sections) and the size (up to 2 mL maximum filling volume) of a microtiter plate well were also considered as influencing parameters. To get an indication of the hydrodynamic behavior of the liquid phase in a well, images were taken during shaking and the liquid height derived as a characteristic parameter. The investigations revealed that the OTR(max) is predominantly dependent on the specific mass transfer area (a) for the considered conditions in round-shaped wells. The mass transfer coefficient (k(L)) in round-shaped wells remains at a nearly constant value of about 0.2 m/h for all shaking intensities, thus within the range reported in the literature for surface-aerated bioreactors. The OTR(max) in round-shaped wells is strongly influenced by the interfacial tension, determined by the surface tension of the medium used and the surface properties of the well material. Up to a specific shaking intensity the liquid surface in the wells remains horizontal and no liquid movement can be observed. This critical shaking intensity must be exceeded to overcome the surface tension and, thus, to increase the liquid height and enlarge the specific mass transfer area. This behavior is solely specific to microtiter plates and has not yet been observed for larger shaking bioreactors such as shaking flasks. In square-shaped microtiter plate wells the corners act as baffles and cause a significant increase of OTR(max), a, and k(L). An OTR(max) of up to 0.15 mol/L/h can be reached in square-shaped wells.

“…Of greater importance is the equilibrium position of the cells at which time the steady-state analysis takes place. The positions of cells after redistribution will not only grade the oxygen consumption rate, but will also present a greater physical blockage to the movement of oxygen through the periphery of the gel because of the lower diffusivity of oxygen through cells than water (Ho and Ju, 1988). Hence, these factors will affect both the diffusivity and reaction terms of the diffusion equation, which have been modified with the following equation of inhomogeneous cell density.…”

Section: Cellular Oxygen Consumptionmentioning

confidence: 99%

Analysis of oxygen transport in a diffusion‐limited model of engineered heart tissue

Brown

MacLellan

Laks

et al. 2007

Cardiac tissue engineering has made notable progress in recent years with the advent of an experimental model based on neonatal cardiomyocytes entrapped in collage gels and purified basement membrane extract, known as "engineered heart tissues" (EHTs). EHTs are a formidable display of tissue-level contractile function and cellular-level differentiation, although they suffer greatly from mass transport limitations due to the high density of metabolically active cells and the diffusion-limited nature of the hydrogel. In this report, a mathematical model was developed to predict oxygen levels inside a one-dimensional, diffusion-limited model of EHT. These predictions were then compared to values measured in corresponding experiments with a hypoxia-sensitive stain (pimonidazole). EHTs were cast between two plastic discs, which allowed for mass transfer with the culture medium to occur in only the radial direction. EHTs were cultured for up to 36 h in the presence of pimonidazole, after which time they were snap-frozen, histologically sectioned, and stained for bound pimonidazole. Quantitative image analysis was performed to measure the distance from the culture medium at which hypoxia first occurs under various conditions. As tested by variation of simple design parameters, the trends in oxygen profiles predicted by the model are in reasonable agreement with those obtained experimentally, although a number of ambiguities related to the specific model parameters led to a general overprediction of oxygen concentrations. Based on the sensitivity analysis in the present study, it is concluded that diffusion-reaction models may offer relatively precise predictions of oxygen concentrations in diffusion-limited tissue constructs.