Mixing, Velocity and Turbulence Characteristics of Shaken Bioreactors

The thesis describes an experimental investigation of the flow in a shaken bioreactor of cylindrical geometry with a flat bottom. Several reactor designs can be distinguished that attain mixing in different ways: oscillatory flow mixers (OFM), static mixers, stirred vessels and shaken bioreactors. Shaken bioreactors are often small-scale mixers (microwells) employed in the early stage of bioprocess development (i.e. microbial fermentation, bioconversion and product recovery techniques), before the developed process is implemented in a large-scale industrial stirred tank. However, despite their wide use, little is known about the fluid mechanics of these systems. In the present study Particle Image Velocimetry (PIV) measurements are carried out to determine the variation of the flow dynamics in a cylindrical shaken geometry for different operating conditions such as medium height, shaking frequency, orbital shaking diameter, cylinder inner diameter and fluid viscosity. In the first part phase-resolved measurements are carried out with PIV to provide a thorough characterisation of the flow and mixing dynamics occurring in a cylindrical shaken bioreactor for a fluid of low viscosity (i.e. water). From this analysis a flow scaling law based on the Froude number, Fr, is identified, which correlates the shape and inclination of the free surface to the occurrence of a flow transition. More specifically it was found that at low Fr the mean flow ischaracterised by a toroidal vortex with its axis aligned along the azimuthal direction, while after flow transition the free surface exhibits a phase lag and a vortical structure with a vertical axis that precesses around the cylinder axis is present. In the second part of the thesis flow characteristics, such as the interfacial area, circulation time, vortex size and location, kinetic energy and viscous dissipation rate of kinetic energy for a fluid of low viscosity are analysed in depth. The free surface interfacial area was directly measured by image analysis to assess oxygen transfer potential and was compared to an analytical solution valid for low Fr. The non-dimensional time and length scales of the vortical structures occurring in the cylindrical bioreactor were determined to provide an insight into the mixing dynamics, while a Reynolds decomposition analysis of the kinetic energy was carried out to assess the onset of a laminar-turbulent flow transition with increasing Fr. Direct measurements of the viscous dissipation rate of the kinetic energy, ǫ, were obtained across the tank to help assess micro-mixing and identify regions in the bioreactor experiencing higher levels of viscous stresses that can potentially affect cell growth. In the third part of the thesis the flow obtained with Newtonian fluids of higher viscosity is investigated and the flow scaling law determined for water is extended to a broader range of viscosity. A flow transition map based on Fr and Re is identified and four main regions characterised by different mean flow dynamics are shown. The turbulent kinetic energy levels and shear rate magnitudes are assessed for different combinations of Fr and Re. The results offer valuable new information for the design of mixing processes and crucial data to validate computational fluid dynamics simulations of cylindrical shaken bioreactors

On the effect of surfactants on drop coalescence at liquid/liquid interfaces

In this work the effect of surfactants on the coalescence of a drop with a flat aqueous-organic interface was experimentally investigated. A high speed Particle Image Velocimetry (PIV) system was used to obtain velocity profiles and kinetic energy per unit mass distribution inside the coalescing droplets. Different mass ratios of surfactant to oil below the CMC value, up to ϕ = 5 × 10−4, of a non-ionic surfactant dissolved in the organic phase were studied. It was found that an increase in the surfactant concentration promoted the deformation of the interface before the film that separated the drop from the interface ruptured. A high surfactant concentration also increased the time needed for film rupture. When rupture occurred, two counter-rotating vortices formed inside the droplet on either side of the rupture point, which moved upwards with time. The propagation of the vortices inside the droplet was faster for low surfactant concentrations, while the intensities of the two counter-rotating vortices significantly decreased for increasing surfactant concentration. At the early stages of coalescence after film rupture, the kinetic energy per unit mass was mainly distributed near the bottom part of the droplet, while at later stages it was distributed near the upper part of the droplet

An experimental study on the drop/interface partial coalescence with surfactants

This paper presents investigations on the partial coalescence of an aqueous drop with an organicaqueous interface with and without surfactants. The organic phase was different silicone oils and the aqueous phase was a glycerol-water solution at different concentrations. It is found that when the surfactant Span 80 is introduced into the organic phase, the partial coalescence region is reduced in the Oh-Bo coalescence map. The range of the inertio-capillary regime reduces when surfactants are present, while the drop size ratio decreases with increasing surfactant concentration. The velocity fields inside the aqueous drop were studied with high speed particle image velocimetry for the first time. In the surfactant-free system, it was found that the inward motion of the fluids at the upper part of the drop favours the generation of a liquid cylinder at the early stages of coalescence. The pressure gradient created by the downward stream at the bottom of the liquid cylinder drives the pinch-off of the secondary drop. When surfactants are present, the rupture of the film between the drop and the interface occurs at an off-axis location. The liquid cylinder formed in this case is not symmetric and does not lead to pinch-off. It is also found that the vortices inside the droplet have little impact on the partial coalescence

Experimental investigation of the solid-liquid separation in a stirred tank owing to viscoelasticity

In this study, we investigated the motion of solid particles dispersed in highly viscous complex fluids agitated in a stirred vessel. We used a refractive index matching method and a combination of planar laser- induced fluorescence (PLIF), particle image velocimetry (PIV), and particle-tracking velocimetry (PTV) techniques to measure the velocity fields of the solid and fluid phases simultaneously along with the spatiotemporal distribution of the solids in the tank. The experimental data show that in a Newtonian ambient fluid, particles disperse uniformly in the plane of measurement, while in a strongly shear-thinning viscoelastic ambient fluid they tend to accumulate in the core of the vortices formed in the flow domain. We found that the solids migrate to the core of the vortices also when the ambient fluid is a Boger fluid, i.e., viscoelastic but not shear thinning. The effect of the first normal stress difference, N1, on the vortex sizes and circulation intensities was also examined, with both properties decreasing for increasing N1. Finally, we observed that the clustering time of the solids in the vessel for viscoelasticity-induced migration was at least three orders of magnitude lower than that obtained from the literature for inertia-induced migration

Laser induced fluorescence studies on the distribution of surfactants during drop/interface coalescence

The spatiotemporal distribution of fluorescent surfactants on the merging interfaces during the coalescence of an aqueous drop with an organic/aqueous flat interface was studied experimentally with high-speed laser induced fluorescence. The aqueous phase was a 46% glycerol solution, while the organic phase was a 5 cSt silicone oil. A fluorescently tagged surfactant was used at a concentration of 0.001 mol/m3 in the aqueous phase. To vary the concentration of surfactants on the interfaces, the drop and the flat interface were left to stand for different times before the coalescence experiments (different interface ages). It was found that when a drop rested on the interface, the surfactants adsorbed on the interfaces were swept outwards by the draining liquid film between the drop and the flat interface and reached a peak value at 0.75Rh away from the centre of the film, where Rh is the horizontal drop radius. After the film rupture, the concentration of the surfactants at the tip of the meniscus increased. Once the film had retracted, the concentration of the surfactants peaked at the meniscus at the bottom of the drop. As the liquid in the drop started to merge with its homophase, the drop formed a cylinder from the upward capillary waves on the drop surface. The surfactant concentration was found to be low at the top of the liquid cylinder as the interface was stretched by the convergence of the capillary waves. Subsequently, the cylinder began to shrink and the top part of the drop acquired a high surfactant concentration

Computational fluid dynamic studies of mixers for highly viscous shear thinning fluids and PIV validation

Agitation of highly viscous shear thinning fluids is normally conducted with complex impeller designs. Often, impellers almost as large as the tanks containing them and impeller blades equipped with holes are adopted in industry. In this work, we studied experimentally the main features of the flow generated by this type of impellers for a mixture of glycerol with a carbomeric gel by means of particle image velocimetry. The experiments were conducted at temperatures ranging from 40 to 60 °C and impeller speeds ranging from 40 to 140 rpm. In all cases, the flow regime was laminar or in the transition region. We also used computational fluid dynamics simulations to describe the behaviour of the mixer, validating the results experimentally with good agreement. We used the numerical results to obtain information on the performance of the mixer, determining the locations and size of vigorous agitation zones and the local effect of the holes present on the impeller blades. The power curves of the mixer were obtained, and the mixer efficiency in terms of power consumption was found to be similar to other impellers used to mix highly viscous non-Newtonian fluids

Mode decomposition and Lagrangian structures of the flow dynamics in orbitally shaken bioreactors

In this study, two mode decomposition techniques were applied and compared to assess the flow dynamics in an orbital shaken bioreactor (OSB) of cylindrical geometry and flat bottom: proper orthogonal decomposition and dynamic mode decomposition. Particle Image Velocimetry (PIV) experiments were carried out for different operating conditions including fluid height, h, and shaker rotational speed, N. A detailed flow analysis is provided for conditions when the fluid and vessel motions are in-phase (Fr = 0.23) and out-of-phase (Fr = 0.47). PIV measurements in vertical and horizontal planes were combined to reconstruct low order models of the full 3D flow and to determine its Finite-Time Lyapunov Exponent (FTLE) within OSBs. The combined results from the mode decomposition and the FTLE fields provide a useful insight into the flow dynamics and Lagrangian coherent structures in OSBs and offer a valuable tool to optimise bioprocess design in terms of mixing and cell suspension. Published by AIP Publishing. https://doi.org/10.1063/1.501630

Surfing of drops on moving liquid–liquid interfaces

The delayed coalescence of drops with the interface between a moving aqueous layer and an oil phase is investigated in a novel flow channel. Drops are released onto oil–aqueous interfaces moving at velocities from 0 cm s−1 up to 3.4 cm s−1 . The evolution of the drop shape, the film thickness between the drop and the bulk liquid, and the velocities of the drop surface and the bulk interface were measured with planar laser-induced fluorescence. As the interface speed increases, the drop coalescence is delayed. This is attributed to the lubrication pressure that develops in the draining film. This pressure was calculated by using the drop shape and the tangential velocities of the drop surface and the bulk interface, and was shown to increase with the interface velocity. The film forming between the drop and the bulk liquid has a dimple shape, symmetric about the centreline. With increasing interface velocity, the dimple shifts to the front part of the drop, resulting locally in a low pressure, which leads to film rupture. As the film breaks, ‘oil drops on a string’ formations are entrained into the water phase, which is rarely seen when a drop coalesces with a stationary liquid–liquid interface. The velocity fields in the drop were investigated with particle image velocimetry. It is found immediately after reaching the interface that the drops accelerate to reach the interface speed. Initially there is a strong internal circulation in the drops, which decays quickly as the drops approach the speed of the interface