Confining mycelial growth to porous microbeads: a novel technique to alter the morphology of non-newtonian mycelial cultures

In an effort to alter the filamentous morphology of Penicillium chrysogenum cells, a technique was developed to confine the growth of the mycelia to porous celite beads. The pore matrix of these beads was found to be very effective for entrapping mycelial cells and spores. The entrapped spores were used to initiate the fermentations in shake flask cultures. Significant increases in final cell densities were obtained in the confined cell cultures reaching up to 60 g/L cells. This is nearly double the cell concentration attainable in free cell cultures grown in the absence of beads. Cell loadings up to 0.55 g cells per bead were obtained in the confined cell cultures. In the later stages of the fermentations, the specific oxygen uptake rates in the confined cell cultures were found to decrease with respect to free cell cultures.     

Hydrodynamics and mass transfer in Aspergillus niger fermentations in bubble column and loop bioreactors

The influence of Aspergillus niger broth rheology, bioreactor geometry, and superficial gas velocity on the volumetric liquid phase oxygen transfer coefficient (k(L)a(L)), riser gas holdup (epsilon(GR)), and circulating liquid velocity (u(LR)) was studied in a bubble column (BC) and two external-circulation-loop airlift (ECLAL) bioreactors. The results are compared to those of previous studies on homogeneous fluids and in particular with a recent study on non-Newtonian carboxymethylcellulose (CMC) solutions conducted in the same contactors used for the A. niger fermentations. As expected from the CMC-based studies, in the heterogeneous broths of A. niger epsilon(GR), k(L)a(L), and u(LR) decreased with increasing broth apparent viscosity; epsilon(GR) and k(L)a(L) decreased with increasing downcomer-to-riser cross-sectional area ratio, A(d)/A(r), whereas u(LR) increased with increasing A(d)/A(r). Gas holdup data in the airlift fermentations of A. niger were well predicted by the CMC-based correlation. However, the CMC-based correlations produced conservative estimations of k(L)a(L) and overestimates of u(LR) compared to the observed values in the A. niger broths.     

Modelling Xanthomonas campestris batch fermentations in a bubble column

Rate and yield expressions relating to biomass and xanthan formation and to nitrogen, glucose, and oxygen consumption were established for Xanthomonas campestris batch fermentations in a bubble column. Microbial growth was described by the logistic rate equation, characterized by a maximum specific growth rate mu(M) = 0.5 h(-1) and a maximum attainable cell concentration provided by nitrogenous compounds. With regard to carbon metabolism, the decrease with time in experimental yields and in the experimental specific rates of xanthan production and glucose assimilation demonstrated the inadequacy of the Luedeking-Piret model. These decreases were connected to the simultaneous drop in dissolved-oxygen tension observed during xanthan synthesis. The knowledge of metabolic pathways and energetic balance were used to establish the relationships between substrate utilization, ATP generation, and xanthan production. The model was structured by assuming the oxygen limitation of both the respiration rate and the efficiency of the oxidative phosphorylation mechanism (P/O ratio). Consequently, the specific rates and yield expressions became dependent on the dissolved-oxygen tension, i.e., of the volumetric oxygen transfer in the fermentor.     

Oxygen mass transfer into aerated CMC solutions in a bubble column

Differing findings on the volumetric mass transfer coefficients k(L)a in CMC solutions in bubble column bioreactors have been reported in the literature. Therefore, oxygen mass transfer was studied again in CMC solutions in a 14-cm-i.d. x 270-cm-height bubble column using different spargers. The k(L)a values were determined along with the dispersion coefficients by fitting the prediction of the axial dispersed plug model with the experimental oxygen concentration profiles in the liquid phase. Surprisingly, the obtained liquid phase dispersion coefficients for CMC solution are higher than one would expect from correlations. The k(L)a data depend largely on the flow regime. In general, they are lower than those reported in the literature. The data for developing slug and established slug flow are dependent on the gas velocity and the effective viscosity of the solution and can br correlated by a simple correlation. This correlation describes k(L)a values measured on fermentation broth of Penicillium chrysogenum with striking agreement.     

Hydrodynamics and mass transfer in miniaturized bubble column bioreactors

Miniaturized bubble columns (MBCs) have different hydrodynamics in comparison with the larger ones, but there is a lack of scientific data on MBCs. Hence, in this study, the effect of gas hold-up, flow regimes, bubble size distribution on volumetric oxygen mass transfer coefficient at different pore size spargers and gas flow rates in MBCs in the presence and absence of microorganisms were investigated. It was found that flow regime transition occurred around low gas flow rates of 1.18 and 0.85 cm/s for small (16–40 µm) and large (40–100 µm) pore size spargers, respectively. Gas hold-up and K_La in MBC with small size sparger were higher than those with larger one, with an increasing effect in the presence of microorganisms. A comparison revealed that the wall effect on the flow regime and gas hold-up in MBCs was greater than bench-scale bubble columns. The K_La values significantly increased up to tenfold using small pore size sparger. In the MBC and stirred tank bioreactors, the maximum obtained cell concentrations were OD₆₀₀ of 41.5 and 43.0, respectively. Furthermore, it was shown that in MBCs, higher K_La and lower turbulency could be achieved at the end of bubbly flow regime.

     

Oxygen mass transfer in a bubble column bioreactor containing lysed yeast suspensions

Summary Liquid-phase volumetric oxygen transfer coefficients were evaluated in a bubble column containing yeast suspensions, using the instationary oxygen absorption method and a polarographic oxygen electrode. The electrode time lag was found to be independent of both the system studied and the operating conditions. The volumetric oxygen mass transfer coefficients k _L a could be reasonably predicted by calculating k _L from the equation derived by Bhavaraju et al. or the empirical equation of Calderbank and Moo-Young and a from the experimental gas hold-up values.Nomenclature a Exponent in Eq.6 or specific gas-liquid interfacial area based on reactor volume m - b Exponent in Eq. 6 - C Constant in Eq 6 or oxygen concentration in the liquid phase g/ml - C ^* Equilibrium oxygen concentration g/ml - C ₀ Oxygen concentration in the liquid phase at t=0 g/ml - C _E Oxygen concentration as determined by the polarographic electrode g/ml - D _B Bubble equivalent diameter mm - D _l Oxygen diffusivity in the liquid phase m²/s - g Acceleration of gravity m/s² - K Consistency index Pasⁿ - K _L Liquid-phase mass transfer coefficient m/s - n Power law exponent - Pe _sw Peclet number based on bubble swarm velocity - S _C Schmidt number - Sh Sherwood number - i Time s - U _B Bubble rise velocity in infinite medium m/s - U _g Superficial air velocity based on column cross-sectional area m/s - U _sw Bubble swarm velocity defined by Eq.15 m/s - Y _MSW Mass transfer coeficient correction factor for mobile interfaces in pseudo-plastic fluids Eq. 7 - Y _MSW Mass transfer coefficient correction factor for immobile interface in pseudo-plastic fluids Eq. 8 Greek letters _l Density of liquid g/ml - _sus Density of unaerated suspension g/ml - _{wet cell} Density of yeast wet cells g/ml - _l Viscosity of the liquid Pas - _app Apparent viscosity of power law fluid Pas - _E Electrode time lag s - _l Time lag due to resistance of the gas-liquid interface s - _g Gas hold-up, volume fraction occupied by the gas phase - _l Liquid hold-up - _c Wet cell volume fraction     

Oxygen mass transfer and gas holdup in a bubble column bioreactor with biosynthesis liquids

The influence of the rheology of some antibiotic biosynthesis liquids produced by Streptomyces aureofaciens, Nocardia mediterranei and Penicillium chrysogenum on the volumetric liquid phase oxygen transfer coefficient, k_La, and gas holdup, ε_G, together with the influence of superficial gas velocity, were studied in a bubble column bioreactor, using samples of fermentation liquids taken from industrial stirred tank fermenters, at 30-hour intervals during fermentation batch. The results were compared to those of previous studies from literature on non-Newtonian homogeneous fluids, such as CMC-Na, xanthan and starch solutions, respectively. In the heterogeneous broths, ε_G and k_La decreased with increasing apparent viscosity of the broth and increased with increasing superficial velocity. The experimental data were correlated using non-linear regression with correlation coefficients above 0.85.     

Influence of antifoam agents on gas hold-up and mass transfer in bubble columns with non-newtonian fluids

Summary Experimental studies on the effect of antifoam agents on the performance of bubble columns with non-Newtonian fluids have been conducted. It is found that the gas hold-up and volumetric mass transfer coefficient in the case of water were reduced due to the addition of antifoam agents. It was found that this decrease in volumetric mass trasfer coefficient is substantial but in the aqueous solutions of polymers the effect becomes weaker as the concentration of polymers becomes higher. When the concentration of polymers became higher than a certain value, the volumetric mass transfer coefficient in the aqueous solutions of polymers with antifoam agents was higher than that without antifoam agents.Nomenclature a Specific surface area, 1/m - D _c Column diameter, m - d _max Diameter of the largest bubble stable against breakup, m - d _min Diameter of the smallest bubble stable against coalescence, m - g Gravitational acceleration, m/s² - H _l Clear liquid height, m - h Rupture thickness of the liquid film, m - K Consistency index in a power-law model, Pa·s ⁿ - k _l Liquid-phase mass transfer coefficient, m/s - n Flow index in a power-law model - u _sg Superficial gas velocity, m/s Greek letters Shear rate, 1/s - Gas hold-up - Energy dissipation per unit mass, m²/s³ - Viscosity, Pa·s - p Density, kg/m³ - Surface tension, N/m - Shear stress-Pa     

Permeabilization of phospholipid bilayer membranes induced by gas-liquid flow in an airlift bubble column

The permeability of 5(6)-carboxyfluorescein (CF) through the phospholipid bilayer membranes was measured by using the system in which the CF-containing phospholipid vesicles (liposomes) were suspended in the gas-liquid flow in an external loop airlift bubble column. The airlift was operated at various superficial gas velocities UG up to 2.4 cm/s at 25 and 40 degrees C. The CF-containing liposomes composed of POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) had the nominal diameters of 50, 100, and 200 nm. The 50- and 100-nm liposomes were stable at 40 degrees C for 5 h even at a high UG value of 2.4 cm/s based on the observed turbidity of the liposome suspension in the airlift. On the other hand, the 200-nm liposomes were stable at a low UG value of 1.4 cm/s, although a progressive decrease in size of the liposomes was implied at the high UG value of 2.4 cm/s. The permeability coefficient PCF of CF through the lipid membrane of the 100-nm liposomes was significantly increased with increasing UG at a high temperature of 40 degrees C, while at a low temperature of 25 degrees C the PCF value was little dependent on UG. As a typical result on the above liposomes, the PCF value (9.2 x 10(-11) cm/s) at 40 degrees C and UG = 2.4 cm/s in the airlift was more than 15 times larger than that at 25 degrees C in the static liquid corresponding to UG = 0. In addition, the dependence of the PCF value on UG at 40 degrees C became more significant with increasing the size of liposomes suspended. The results obtained revealed that the permeability of the liposome membranes could be regulated by suspending the liposomes in the gas-liquid flow in the airlift without modulating the membrane composition of liposomes.     

Gas holdup and mass transfer characteristics of carboxymethyl cellulose solutions in a bubble column with a radial gas sparger

The gas phase holdup and mass transfer characteristics of carboxymethyl cellulose (CMC) solutions in a bubble column having a radial gas sparger have been determined and a new flow regime map has been proposed. The gas holdup increases with gas velocity in the bubbly flow regime, decreases in the churn-turbulent flow regime, and increases again in the slug flow regime. The volumetric mass transfer coefficient (k _La) significantly decreases with increasing liquid viscosity. The gas holdup and k _La values in the present bubble column of CMC solutions are found to be much higher than those in bubble columns or external-loop airlift columns with a plate-type sparger. The obtained gas phase holdup ( _g) and k _La data have been correlated with pertinent dimensionless groups in both the bubbly and the churn-turbulent flow regimes.List of Symbols a m^–1 specific gas-liquid interfacial area per total volume - A _d m² cross-sectional area of downcomer - A _r m² cross-sectional area of riser - d _b m individual bubble diameter - d _vs m Sauter mean bubble diameter - D _c m column diameter - D _L m²/s oxygen diffusivity in the liquid - Fr Froude number, U _g/(g D_c)^1/2 - g m/s² gravitational acceleration - G _a Galileo number, gD _c ^{3 2}/² _app - H _a m aerated liquid height - H _c m unaerated liquid height - K Pa · sⁿ fluid consistency index - k _L a s^–1 volumetric mass transfer coefficient - n flow behavior index - N _i number of bubbles having diameter d _bi - Sc Schmidt number, _app/( D _L) - Sh Sherwood number, k _L a D _c ² /D_L - U _sg m/s superficial gas velocity - U _gr m/s superficial riser gas velocity - V _a m³ aerated liquid volume - V _c m³ unaerated liquid volume - N/m surface tension of the liquid phase - _g gas holdup - _app Pa · s effective viscosity of non-Newtonian liquid - kg/m³ liquid density - ý s^–1 shear rate - Pa shear stress     

Simulation of algae growth in a bench-scale bubble column reactor

The growth of the marine red microalga Porphyridium sp. in a bubble-column photobioreactor was simulated. The proposed model constitutes a dynamic integration of the kinetics of photosynthesis and photoinhibition with the fluid dynamics of the bubble column, including the effects of shear stress on the kinetics of growth. The kinetic data used in the model were obtained in independent experiments run in a thin-film photobioreactor with defined light/dark cycles. The maintenance term was modified to take into account the effects of liquid flow in the bioreactor on the growth rate. A hybrid method proposed for the approximate solution of the equations gave an appreciable reduction of the calculation time. Extrapolations of the model indicated the possibility of predicting the optimal diameter for an assembly of bubble column photobioreactors. Satisfactory fit was found with the experimental results of biomass growth in a 13-liter bubble column.     

Oxygen transfer and mycelial growth in a tubular loop fermentor

A 22 m long. 20 liter tubular loop fermentor (TLF) has been tested for oxygen transfer characteristics and as a reactor for mycelial growth. Model calculations show that the flow pressure drop has an important influence on the axial oxygen profiles. A design model that accounts for this influence is presented. Using the model, K_L a values are calculated from the results of sulfite oxidation experiments. These are correlated with power consumption and aeration rates. The K_L a dependence on aeration rate was found to be less than found with tank reactors. The growth kinetics of three metabolite-producing mycelial organisms in the TLF are presented: a Streptomyces, a Fusarium, and a Acrophialophora. In order to determine the influence of reactor type on the growth and product formation, these cultures have been grown in tanks and shake flasks. The antibiotic, product spectrum of Streptomyces is compared on the basis of inhibition tests and it is shown that the distribution of products is reactor dependent. The Fusarium culture produced a previously unknown metabolite, whose concentration in the loop fermentor was four times higher than in a shake flask. The Acrophialophora culture grew twice as fast in the loop fermentor, but produced essentially none of the specific product. Power Consumptions of up to 8 kW/m³ in the tubular fermentor did not appear to harm the mycelia.     

The influence of polymeric membrane gas spargers on hydrodynamics and mass transfer in bubble column bioreactors

Gas sparging performances of a flat sheet and tubular polymeric membranes were investigated in 3.1 m bubble column bioreactor operated in a semi batch mode. Air–water and air–CMC (Carboxymethyl cellulose) solutions of 0.5, 0.75 and 1.0 % w/w were used as interacting gas–liquid mediums. CMC solutions were employed in the study to simulate rheological properties of bioreactor broth. Gas holdup, bubble size distribution, interfacial area and gas–liquid mass transfer were studied in the homogeneous bubbly flow hydrodynamic regime with superficial gas velocity (U _G) range of 0.0004–0.0025 m/s. The study indicated that the tubular membrane sparger produced the highest gas holdup and densely populated fine bubbles with narrow size distribution. An increase in liquid viscosity promoted a shift in bubble size distribution to large stable bubbles and smaller specific interfacial area. The tubular membrane sparger achieved greater interfacial area and an enhanced overall mass transfer coefficient (K _La) by a factor of 1.2–1.9 compared to the flat sheet membrane.     

Hydrodynamic characteristics and gas-liquid mass transfer in a biofilm airlift suspension reactor

The hydrodynamics and mass transfer, specifically the effects of gas velocity and the presence and type of solids on the gas hold-up and volumetric mass transfer coefficient, were studied on a lab-scale airlift reactor with internal draft tube. Basalt particles and biofilm-coated particles were used as solid phase. Three distinct flow regimes were observed with increasing gas flow rate. The influence of the solid phase on the hydrodynamics was a peculiar characteristic of the regimes. The volumetric mass transfer coefficient was found to decrease with increasing solid loading and particle size. This could be predominantly related to the influence that the solid has on gas hold-up. The ratio between gas hold-up and volumetric mass transfer coefficient was found to be independent of solid loading, size, or density, and it was proven that the presence of solids in airlift reactors lowers the number of gas bubbles without changing their size. To evaluate scale effects, experimental results were compared with theoretical and empirical models proposed for similar systems.     

Characterization of gas transfer and mixing in a bubble column equipped with a rubber membrane diffuser

Gas transfer and mixing were characterized in a 32-L bubble column reactor equipped with a commercially available rubber membrane diffuser. The performance of the membrane diffuser indicates that the slits in the membrane are best described as holes with elastic lids, acting as valves cutting off bubbles from the gas stream. The membrane diffuser thus functions as a one-way valve preventing backflow of liquid. Our design of the bottom plate of the reactor enabled us to optimize the aeration by changing the tension of the membrane. We thereby achieved mass transfer coefficients higher than those previously reported in bubble columns. A strong dependence of mass transfer on gas holdup and bubble size was indicated by estimates based on these two variables. The microalga, Rhodomonas sp. , sensitive to chemical and physical stress, was maintained for 8 months in continuous culture with a productivity identical to cultures grown in stirred tank reactors. Copyright 1998 John Wiley & Sons, Inc.     

Characterization of factors influencing the growth of Anabaena variabilis in a bubble column reactor

The combined effect of superficial gas velocity, pH, initial phosphate concentration, and light intensity on cell growth was investigated for the mass production of cyanobacterial cells. The light intensity was manipulated to maintain a specific irradiation rate (q(i)) at a constant level for high cell density culture. The optimum condition for the batch culture was achieved at a superficial gas velocity of 2.0 cm/s, pH 7.0, and an initial phosphate concentration of 55 mg/l when the specific irradiation rate was controlled above 11.5 micromol/s/g dry cell. In this condition, the specific growth rate and cell productivity were 1.47 day(-1) and 0.98 g dry cell/l/day, respectively.     

Measuring carbon monoxide gas-liquid mass transfer in a stirred tank reactor for syngas fermentation

Carbon monoxide-liquid mass transfer rates in a water-filled stirred tank reactor are determined using a myoglobin protein method to measure dissolved carbon monoxide concentrations as a function of time. Data are acquired over a range of stirrer speeds (200 < or = N < or = 600 rpm) and gas flow rates (1 < or = Q < or = 6 L/min), corresponding to a gas retention time range of 1.2-7 min. Volumetric CO-water mass transfer coefficients range from 0.003 to 0.043 s(-1) and are well-correlated using the power density and the superficial gas velocity.     

Production of proteolytic enzyme by Rhizopus oryzae in a bubble column bioreactor

Production of proteolytic enzyme in a bubble column bioreactor was carried out using the microorganism Rhizopus oryzae. Optimization of the production conditions were done adopting the EVOP (evolutionary operation technique). Studies on two variable and three variable parameters were carried out. The variable parameters were air flow rate, aeration time and incubation period. 8 hours aeration time per day, 1.66 vvm air flow rate and 2 days incubation was found to be optimum.     

Thienamycin production by immobilized cells of Streptomyces cattleya in a bubble column

A novel 2.0-L columnar reactor has been developed for the production of thienamycin by cells of Streptomyces cattleya attached to celite particles. Successful immobilization of cells was achieved by operating the column continuously at a high dilution rate during the growth phase. Scanning electron micrographs of the celite particles indicate the involvement of subcellular fibrils in the attachment of cells to the solid surfaces. Reactor operation was divided into two distinct phases-a growth phase and a production phase. The kinetics of attached growth and thienamycin production were found to be strongly influenced by nutrient concentrations. The influences of nutrient concentration on CO(2) production and thienamycin production during both the growth phase and the production phase are discussed.     

Measurements on the interfacial areas of hydrocarbon in yeast fermentations and relationships to specific growth rates

The transport of insoluble substrates such as hydrocarbons to microorganisms is often postulated to be dictated by the availability of the hydrocarbon surface area. Many publications, qualitative and quantitative, have appeared to substantiate this hypothesis. Experiments have been performed in our laboratory to assess the absolute values of the interfacial area of hexadecane as the carbon source for the growth of Candida intermedia. A sedimentometer, mounted directly in the fermentor, was used to measure the interfacial hydrocarbon area during active growth of this organism. The specific hydrocarbon interfacial area was found to be directly related to the impeller speed, hydrocarbon concentration and surfactant concentration in a 1-liter working volume, turbine-agitated fermentor. The specific growth rate was in turn found to be directly related to the specific hydrocarbon interfacial area. Lastly, cessation of logarithmic growth and onset of linear growth was found at all instances to be governed by the specific hydrocarbon surface area.