Characterization of a pilot plant airlift tower loop reactor: III. Evaluation of local properties of the dispersed gas phase during yeast cultivation and in model media

The local properties of the dispersed gas phase (gasholdup, bubble diamater, and bubble velocity) were measured and evaluated at different positions in the riser and downcomer of a pilot plant reactor and, for comparison, in a laboratory reactor. These were described in Parts I and II of this series of articles during yeast cultivation and with model media. In the riser of the pilot plant reactor, the local gas holdup and bubble velocities varied only slightly in axial direction. The gas holdup increased considerably, while the bubble velocity increased only slightly with aeration rate. The bubble size diminished with increasing distance from the aerator in the riser, since the primary bubble size was larger than the equilibrium bubble size. In the downcomer, the mean bubble size was smaller than in the riser. The mean bubble size varied only slightly, the bubble velocity was accelerated, and the gas holdup decreased from top to bottom in the downcomer. In pilot plant at constant aeration rate, the properties of the dispersed phase were nearly constant during the batch cultivation, i.e., they depended only slightly on the cell concentration. In the laboratory reactor, the mean bubble sizes were much larger than in the pilot plant reactor. In the laboratory reactor, the bubble velocities in the riser and downcomer increased, and the mean gas holdup and bubble diameter in the downcomer remained constant as the aeration rate was increased.     

Investigation of the structure of two-phase flow model-media in bubble-column bioreactors

Summary By applying photographic, electrical conductivity, and electrooptical methods, the transverse variation of bubble size and velocity, the local gas holdup, and the local specific gas/liquid interfacial area were estimated in a bench scale bubble-column bioreactor containing distilled water. The liquid velocity profile, the transverse turbulence intensity variations, and the turbulence energy dissipation scale were also measured by a hot film turbulence probe and constant temperature anemometer technique.     

Investigation of the structure of two-phase flow model-media in bubble-column bioreactors

Summary By applying photographic, electrical conductivity, and electrooptical methods, the transverse variation of bubble size and velocity, the local gas hold up, and the local specific gas/liquid interfacial area were estimated in a bench scale bubble-column bioreactor containing model cultivation media. The liquid velocity profile, the transverse turbulence intensity variations, and the turbulence energy dissipation scale were also measured by a hot film turbulence probe and constant temperature anemometer technique.A significant relationship was found between the two-phase flow fluid dynamical state and the transverse variation of the various properties.Symbols M mass - L length - T time - a gas/liquid interfacial area L² - specific gas/liquid interfacial area with regard to the bubbling layer volume L^–1 - D transverse coordinate (measured from the wall of the column) L - d bubble diameter L - d mean bubble diameter L - d_e dynamic equilibrium (maximum stable) bubble diameter L - d_p primary bubble diameter L - d_s Sauter bubble diameter L - E specific energy dissipation rate with regard to the volume of the liquid ML^–1T^–3 - EV_L energy dissipation rate ML²T^–3 - , since =1 g/cm³, E has the same numerical value as E. Therefore, the symbol E is used everywhere in the present paper for E for simplicity and called energy dissipation rate (S.s^–2=Stokes.s^–2) L²T^–3 - E_G or local relative gas holdup - f (r) cross correlation function - g acceleration of gravity LT^–2 - h longitudinal distance from the aerator L - relative turbulence intensity - N_O number of u and crossings T^–1 - n_B bubble frequency T^–1 - r distance between two points 1 and 2 of the cross correlation function L - t time - u instantaneous liquid velocity LT^–1 - mean liquid velocity LT^–1 - mean square fluctuation velocity L²T^–2 - turbulence intensity LT^–1 - w_SG superficial gas velocity LT^–1 - w_SL superficial liquid velocity LT^–1 - or E_G local relative gas holdup LT^–1 - energy dissipation scale L - kinematic liquid viscosity L²T^–1 - liquid density M L^–3 - surface tension M T^–2 - dynamic turbulence pressure M L^–1T^–2 Indices p primary (at the aerator) - e equilibrium (far from the aerator)     

Methods for measuring the properties of the turbulence in bubble flow

Summary A constant temperature hot film anemometer has been used to evaluate mean liquid flow velocity, bubble frequency, turbulence scale and intensity, and the rate of energy dissipation by liquid phase bubble flow.Symbols M mass - L lenght - T time - a gas/liquid interfacial area L² - a=a/V_L specific gas/liquid interfacial area with regard to the volume of the liquid L^–1 - d bubble diameter L - d mean bubble diameter L - d_e dynamic equilibrium (maximum stable) bubble size L - d_p primary bubble diameter L - d_s Sauter bubble diameter L - E specific energy dissipation rate with regard to the volume of the liquid ML^–1T^–3 - E V_L energy dissipation rate ML²T^–3 - E=E/ since =1 g cm^–3, E has the same numerical value as E. Therefore, the symbol E is used everywhere in the present paper for E and called energy dissipation rate (S. s^–2=Stokes. s^–2) L²T^–3 - E_G or _G local relative gas hold up L²T^–3 - f() autocorrelation function [Eq. (10)] L²T^–3 - f(r) cross correlation function [Eq. (11)] L²T^–3 - g acceleration of gravity LT^–2 - k constant LT^–2 - k_L mass transfer coefficient LT^–1 - k_La volumetric mass transfer coefficient with regard to the volume of the liquid T^–1 - N₀ number of crossings of u and T^–1 - n_B bubble frequency T^–1 - r distance between two points 1 and 2 of the cross correlation function L - t time T - u momentaneous liquid velocity LT^–1 - mean liquid velocity LT^–1 - mean square fluctuation velocity L²T^–2 - intensity of turbulence LT^–1 - x position coordinate L - V volume of the bubbling layer in the column L³ - V_L volume of the bubble free layer in the column L³ - V electrical voltage (in Fig. 2) L³ - v velocity scale [Eq. (6)] LT^–1 - We_crit critical Weber number [Eq. (4)] LT^–1 - w_SG superficial gas velocity LT^–1 - w_SL superficial liquid velocity LT^–1 - _G or E_G local relative gas hold up LT^–1 - smallest scale [Eq. (6)] L - time delay in the autocorrelation function [Eq. (10)] T - energy dissipation scale [E. (15)] L - _f: Taylor's vorticity scale [E. (14)] L - kinematic viscosity of the liquid L²T^–1 - density of the liquid ML^–3 - surface tension MT^–2 - dynamic pressure of the turbulence [Eq. (8)] ML^–1T^–2 - p primary (at the aerator) - e equilibrium (far from the aerator)     

Investigation of the structure of two-phase flows. Model media in bubble-column bioreactors

Summary The following two-phase properties were evaluated in bubble column reactors with porous plate (5 m pore diameter) or perforated plate (1 mm and/or 3 mm hole diameter) gas distributors using distilled water or a 1% methanol solution: transverse profiles of the mean and Sauter bubble diameters, local gas holdups, true mean liquid and bubble velocities. Furthermore, swarm bubble velocity distributions were evaluted and compared with calculated values.     

Understanding the effect of high gas entrance velocity on Chinese hamster ovary (CHO) cell culture performance and its implications on bioreactor scale-up and sparger design

There are three main potential sources for cell shear damage existing in stirred tank bioreactors. One is the potential high energy dissipation in the immediate impeller zones; another from small gas bubble burst; and third is from high gas entrance velocity (GEV) emitting from the sparger. While the first two have been thoroughly addressed for the scale-up of Chinese hamster ovary (CHO) cell culture knowing that a wide tolerable agitation range with non-damaging energy dissipation exists and the use of shear protectants like Pluronic F68 guard against cell damage caused by bubble burst, GEV remains a potential scale-up problem across scales for the drilled hole or open pipe sparger designs. GEV as high as 170 m/s due to high gas flow rates and relatively small sparger hole diameters was observed to be significantly detrimental to cell culture performance in a 12,000 L bioreactor when compared to a satellite 2 L bioreactor run with GEV of <1 m/s. Small scale study of GEV as high as 265 m/s confirmed this. Based on the results of this study, a critical GEV of >60 m/s for CHO cells is proposed, whereas previously 30 m/s has been reported for NS0 cells by Zhu, Cuenca, Zhou, and Varma (2008. Biotechnol. Bioeng., 101, 751–760). Implementation of new large scale spargers with larger diameter and more holes lowered GEV and helped improve the cell culture performance, closing the scale-up gap. Design of such new spargers was even more critical when hole plugging was discovered during large scale cultivation hence exacerbating the GEV impact. Furthermore, development of a scale down model based on mimicry of the large scale GEV profile as a function of time was proven to be beneficial for reproducing large scale results.     

Absorption of oxygen in highly viscous newtonian and non-Newtonian fermentation model media in bubble column bioreactors

Summary Mean relative gas holdup, slip velocity, bubble size distribution, mean specific interfacial area, and volumetric mass transfer coefficient of oxygen were estimated in sparged columns 14 cm in diameter and 380 and/or 390 cm high with two different aerator types (porous plate and injector nozzle) in highly viscous Newtonian (glycerol solutions) and non-Newtonian (CMC solutions) fluids.For the Newtonian liquids the above properties were estimated as function of the viscosity of the liquid. For the non-Newtonian liquids they were determined as function of the fluid consistency index and flow behavior index. Significant differences between Newtonian and non-Newtonian systems appear. In Newtonian medium k_L a drops with increasing viscosity and already approaches a constant value at =40 cP. In pseudoplastic medium k_L a varies with the fluid consistency and flow behavior indexes in the entire investigated range.In both of these systems the primary bubble population changes into two or three populations along the reactor: the medium bubbles gradually disappear and small and large bubbles are formed.     

Alfvén wave spectrum in a transversely inhomogeneous plasma with dissipation

The spectrum of Alfvén eigenmodes in a transversely inhomogeneous plasma with Ohmic dissipation is studied in the one-fluid MHD approximation. It is established that, along with a discrete spectrum of the modes confined to the plasma boundaries or the extremes of the Alfvén velocity, there always exists a continuous spectrum of aperiodically damped modes, including those with arbitrarily slow damping rates. It is shown that the set of eigenmodes is complete.     

Characterization of a pilot plant airlift tower loop bioreactor. I: evaluation of the phase properties with model media

Investigations were carried out in a 9 m high, 4 m(3) volume, pilot plant airlift tower loop bioreactor with a draft tube. The reactor was characterized by measuring residence time distributions of the gas phase using pseudostochastic tracer signals and a mass spectrometer and by evaluating the mixing in the liquid phase with single-pulse tracer inputs. The local gas holdup and the bubble size (piercing length) were measured with two-channel electrical conductivity probes. The mean residence times and the intensities of the axial mixing in the riser and downcomer and the circulation times of the phases as well as the fraction of the recirculated gas phase were evaluated. The gas holdup in the riser is nearly uniform along the reactor. In the downcomer, it diminishes from top to bottom. The liquid phase dispersion coefficients, D(L), are smaller than those measured in the corresponding bubble columns. In the pilot plant with tap water the following relationship was found: D(Lr) = cw(SG) (n); with c = 203.4; n = 0.5;D(Lr)(cm(2) s(-1);) and W(SG)(cm s(-1)) where D(Lr) is the longitudinal dispersion coefficient in the riser and W(SG) is the superficial gas velocity. The gas phase dispersion coefficients in the riser of the pilot plant, D(Gr), are also enlarged with increasing superficial gas velocity, W(SG), however, no simple relationship exists. Parameter D(Gr) is the highest in the presence of antifoam agents, intermediate in tap water, and the smallest in ethanol solution.     

Components of ecosystem evaporation in a temperate coniferous rainforest, with canopy transpiration scaled using sapwood density

Here we develop and test a method to scale sap velocity measurements from individual trees to canopy transpiration (E(c)) in a low-productivity, old-growth rainforest dominated by the conifer Dacrydium cupressinum. Further, E(c) as a component of the ecosystem water balance is quantified in relation to forest floor evaporation rates and measurements of ecosystem evaporation using eddy covariance (E(eco)) in conditions when the canopy was dry and partly wet. Thermal dissipation probes were used to measure sap velocity of individual trees, and scaled to transpiration at the canopy level by dividing trees into classes based on sapwood density and canopy position (sheltered or exposed). When compared with ecosystem eddy covariance measurements, E(c) accounted for 51% of E(eco) on dry days, and 22% of E(eco) on wet days. Low transpiration rates, and significant contributions to E(eco) from wet canopy evaporation and understorey transpiration (35%) and forest floor evaporation (25%), were attributable to the unique characteristics of the forest: in particular, high rainfall, low leaf area index, low stomatal conductance and low productivity associated with severe nutrient limitation.     

Cell death in the thin films of bursting bubbles.

A sparged gas bubble floating at the liquid interface has a liquid film which drains and thins until the film spontaneously ruptures at a point. This causes rapid retraction of the film, forming a rim of collected fluid. This rim moves at a constant velocity of about 3 m/s and any cells in the bubble film are rapidly accelerated to this velocity in the moving rim. Half of the surface energy originally in the thin film is converted to kinetic energy of the rim, while the rest is dissipated in this rim. The rate of energy dissipation per mass of rim fluid is approximately 9000 m2/s3, which corresponds to a Kolmogorov eddy size of 3.2 microns in fully developed turbulence or a shear stress of 95 N/m2 in laminar flow. Either of these limiting cases presents an environment in which rapid cell death would be expected. Experiments with Sf-9 insect cells suggest that the cell concentration in these thin films is 0.6 times the bulk liquid concentration and that about 20% of these cells are killed when the film ruptures. An equation based on this mechanism accurately predicts the death rate.     

Effects of dissipation and external fields on the dynamics of conformational distortions in DNA

The dynamics of conformational distortions in DNA has been studied in the framework of the simple mathematical model based on the sine-Gordon equation with two additional terms. The first of the terms describes the effects of dissipation, and the second takes the action of external field into account. With the help of the energetic method, an analytical expression for the velocity of local conformational distortion as a function of time has been found, and conditions have been determined under which the influence of dissipation and constant external force are in a balance, providing the distortion to move with a constant velocity along the DNA. The graphs of changes in the velocity of movement of local conformational distortions in different homogeneous polynucleotide chains have been constructed using the model values of the parameters v0 = 189 (m/s), beta(DNA) = 4.25 x 10(-34) (J x s) and F0(DNA) = 3.12 x 10(-22) (J), which determine the initial velocity of the distortion, the coefficient of dissipation, and the value of the external generalized force, respectively. The accordance of the model values used with the experimental values available from the literature is discussed.     

Protein crystallization in stirred systems—scale‐up via the maximum local energy dissipation

Macromolecular bioproducts like therapeutic proteins have usually been crystallized with µL‐scale vapor diffusion experiments for structure determination by X‐ray diffraction. Little systematic know‐how exists for technical‐scale protein crystallization in stirred vessels. In this study, the Fab‐fragment of the therapeutic antibody Canakinumab was successfully crystallized in a stirred‐tank reactor on a 6 mL‐scale. A four times faster onset of crystallization of the Fab‐fragment was observed compared to the non‐agitated 10 µL‐scale. Further studies on a liter‐scale with lysozyme confirmed this effect. A 10 times faster onset of crystallization was observed in this case at an optimum stirrer speed. Commonly suggested scale‐up criteria (i.e., minimum stirrer speed to keep the protein crystals in suspension or constant impeller tip speed) were shown not to be successful. Therefore, the criterion of constant maximum local energy dissipation was applied for scale‐up of the stirred crystallization process for the first time. The maximum local energy dissipation was estimated by measuring the drop size distribution of an oil/surfactant/water emulsion in stirred‐tank reactors on a 6 mL‐, 100 mL‐, and 1 L‐scale. A comparable crystallization behavior was achieved in all stirred‐tank reactors when the maximum local energy dissipation was kept constant for scale‐up. A maximum local energy dissipation of 2.2 W kg^?1 was identified to be the optimum for lysozyme crystallization at all scales under study. Biotechnol. Bioeng. 2013; 110: 1956–1963. © 2013 Wiley Periodicals, Inc.     

Investigation of the structure of two-phase flow fermentation model media in bubble-column bioreactors

Summary Electrical conductivity microprobes have been used to estimate the transverse variation of bubble size, local gas holdup and local specific gas/liquid interfacial area in bench scale bubble column bioreactors containing fermentation model media. Inserted O₂-electrodes and plane parallel windows alter the structure of the two phase flow. Even slight tilting of the column strongly influences the transverse profiles of the bubble size and local gas holdup. The larger bubbles are collected at the wall, where they can be redispersed. These observations open up new possibilities for the construction of bubble column bioreactors.     

Enhanced uptake of dissolved oxygen and glucose by <Emphasis Type="Italic">Escherichia coli</Emphasis> in a turbulent flow

Laboratory experiments were conducted to study the effect of turbulence on Escherichia coli cells in an oscillating grid reactor under conditions of no oxygen transfer to the liquid phase. Fluid flow was quantified at a submillimeter resolution using a particle image velocimetry measuring technique. The root-mean-square estimates of the velocity gradient tensor components indicated the dominance of shear rate deformation in the fluid surrounding E. coli. The E. coli growth rate, dissolved oxygen (DO), and glucose uptake rates were facilitated by fluid-flow energy dissipation in the turbulent fluid. The Kolmogorov length scale (eta(K)) and velocity (u( K )) underlined characteristic scales at which enhanced DO and glucose uptake by E. coli were determined in a turbulent flow in comparison to still-water controls. A first-order power-law relation between the mass transport to the cells and the moving fluid is developed. The combined effects of the enhanced rate of strain at eta(K) scale and uniform velocity at u(K) determined the facilitated DO and glucose fluxes to E. coli. The mass transport to the E. coli was modeled by the Sherwood (Sh)-Péclet (Pe) number relationship by Sh = 1 + 1.08Pe(uK)(0.62) where Pe(uK) is the Péclet number defined by the u(K) velocity scale. The proposed first-order model described experimental data fairly well.     

Different bioreactor configurations for the decolourisation of the azo dye reactive black 5 by Geotrichum sp. CCMI 1019

Decolourisation of the azo dye Reactive Black 5 by Geotrichum sp. CCMI 1019 was studied using stirred tank reactors (STR) and two types of bubble columns (porous plate (PP) bubble column and aeration tube (AT) bubble column). For the bubble columns, the k_La increased with the gas fractional hold-up (ε_G) and the aeration rate. A linear relationship between ε_G and superficial gas velocity was obtained for all reactors. At same aeration rates, the PP bubble columns showed higher k_La and hold-up values than the AT bubble column. In the STRs, large and dense aggregates were formed which adhered to surfaces whereas bubble columns gave smaller and less compact pellets.

Manganese peroxidase and laccase were detected in the extracellular media in all reactors. However, laccase was only detected after the onset of decolourisation, suggesting that additional enzymes may be involved. Mn peroxidase activity was detected (about 46 U/ml) in both the STRs and AT bubble columns but higher values (110 U/ml) were obtained with the PP bubble columns.

Out of the three reactor systems studied, the AT bubble columns gave the most favourable results for Reactive Black 5 decolourisation. Rapid and complete colour removal was obtained throughout the visible spectrum. Bubble columns are simple in design as well as operation and may be useful for the bioremediation of textile wastewater.     

Local gas hydrodynamics in an external-loop airlift reactor: newtonian and non-newtonian fluids

Measurements of local gas phase characteristics are obtained in an external-loop airlift reactor filled with newtonian or viscous non-newtonian liquids. A double-optical fiber probe technique is used. It allows the determination of the axial and radial profiles of gas hold-up, bubbling frequency, bubble size and velocity. In the case of air-water system, the results show a strong effect of radial liquid velocity variation on the gas flow characteristics at the bottom of the riser. In the case of highly viscous non-newtonian solution, the gas flow is strongly affected by the gas distribution just above the gas sparger. This study also points out the bubble coalescence and the break-up phenomena in different liquids and levels in the reactor. Furthermore, the local measurements of bubble size and velocity allows to gain more detailed information on the dynamics of the bubble-flow and shows a tendency of large bubbles to circulate in the column center.     

Fluiddynamic characterization of airlift tower loop bioreactors with and without motionless mixtures

In a 60 l airlift tower reactor with outer loop fluiddynamical measurements were carried out in presence of three motionless mixer modules (Type SMV, Sulzer) in water, 0.6%, 0.9% and 1.2% CMC solutions. The global mixing properties were determined in the liquid and gas phases by tracers. Detailed local measurements revealed differences of local flow patterns and mixing properties in the unhindered riser and in the immediate wake of the mixer module. The local liquid velocities were measured by the pseudorandom heat pulse technique. The local bubble velocities were determined by the ultrasound Doppler technique. The dependence of liquid velocity, gas velocity and gas holdup on the superficial gas velocity were determined. The radial profiles of the mean liquid velocities and their standard deviations were evaluated in water and CMC solutions upstream and downstream of the motionless mixer modules. The radial profiles of the mean large-bubble velocities and mean small-bubble velocities and their standard deviations were determined as well. The influence of the presence of the motionless mixers in the riser on these properties was evaluated.List of Symbols Bo _L w _L L/D _L liquid Bo number - Bo _G w _G L/D _G ,gas Bo number - c tracer concentration - CMC carboxymelthylcellulose - D _G gas dispersion coefficient - D ₁ local liquid dispersion coefficient - D _L liquid dispersion coefficient - D _r riser diameter - d distance between transmitter and detector of the heat pulse probe - E _G gas holdup - HBV horizontal bubble velocity - HLBV horizontal large-bubble velocity - HSBV horizontal small-bubble velocity - L length of the column - l relative distance of the detector position from the tracer injection with respect to L - LBV large-bubble velocity - n number of circulations - OHBV overall horizontal bubble velocity - OVBV overall vertical bubble velocity - P power input - Pe ₁ w ₁d/D₁, local liquid Peclet number - SBV small-bubble-velocity - V _L liquid volume - VBV vertical bubble velocity - VLBV vertical large-bubble velocity - VSBV vertical small-bubble velocity - w _G gas velocity - w ₁ local liquid velocity - w _L liquid velocity - w _SG superficial gas velocity - w _SL superficial liquid velocity - mean residence time of the liquid in the riser H.M.R. thank the Verband der Chemischen Industrie for a Fund der Chemie scholarship     

Influence of dissipation and external field on the dynamics of local deformations in DNA

The dynamics of local deformations in DNA is studied in a simple mathematical model based on the sine-Gordon equation with two additional terms, one accounting for the effects of dissipation and the other for the action of an external field. The energy method is used to derive equations for the velocity of the local deformation as a function of time. Conditions are determined when dissipation and continuous external action are balanced, whereby the deformation can move along the DNA at a constant velocity. The velocity vs. time plots are shown for homopolynucleotide chains at model parameter values: initial velocity v ₀ = 189 m/s, dissipation coefficient \(\overline \beta ^{DNA} \) = 4.25·10^?34 J s, and generalized external force F ₀ ^DNA = = 3.12·10^?22 J. The correspondence of these value with available experimental data is discussed.     

Interactions between the photosystem II subunit PsbS and xanthophylls studied in vivo and in vitro

The photosystem II subunit PsbS is essential for excess energy dissipation (qE); however, both lutein and zeaxanthin are needed for its full activation. Based on previous work, two models can be proposed in which PsbS is either 1) the gene product where the quenching activity is located or 2) a proton-sensing trigger that activates the quencher molecules. The first hypothesis requires xanthophyll binding to two PsbS-binding sites, each activated by the protonation of a dicyclohexylcarbodiimide-binding lumen-exposed glutamic acid residue. To assess the existence and properties of these xanthophyll-binding sites, PsbS point mutants on each of the two Glu residues PsbS E122Q and PsbS E226Q were crossed with the npq1/npq4 and lut2/npq4 mutants lacking zeaxanthin and lutein, respectively. Double mutants E122Q/npq1 and E226Q/npq1 had no qE, whereas E122Q/lut2 and E226Q/lut2 showed a strong qE reduction with respect to both lut2 and single glutamate mutants. These findings exclude a specific interaction between lutein or zeaxanthin and a dicyclohexylcarbodiimide-binding site and suggest that the dependence of nonphotochemical quenching on xanthophyll composition is not due to pigment binding to PsbS. To verify, in vitro, the capacity of xanthophylls to bind PsbS, we have produced recombinant PsbS refolded with purified pigments and shown that Raman signals, previously attributed to PsbS-zeaxanthin interactions, are in fact due to xanthophyll aggregation. We conclude that the xanthophyll dependence of qE is not due to PsbS but to other pigment-binding proteins, probably of the Lhcb type.