Regular articleSparger design as key parameter to define shear conditions in pneumatic bioreactors
Graphical abstract
Introduction
The selection of a bioreactor model for application in an aerobic bioprocess should consider not only the oxygen transfer capability, but also the shear environment, since biochemical processes are extremely sensitive to the shear intensity when fragile animal cells, plant cells, and filamentous microorganisms are used [1]. Excessive shear can cause cell damage, leading to viability loss and cell disruption or disintegration, so bioreactors should provide moderate or low shear environments, in order to avoid such effects [1,2].
The traditional parameter applied for evaluation of shear conditions is the average shear rate (), which is extensively used for comparison of the performances of different bioreactor models. Several studies have evaluated this parameter, with the development of methodologies and correlations for bioreactor types including the bubble column [[3], [4], [5], [6], [7]], airlift [2,6,[8], [9], [10], [11]], and stirred tank [7,[12], [13], [14], [15], [16]]. Based on this approach, the effect of on microbial growth, enzyme activity, and biomolecule production has been evaluated [17]. A general observation is that pneumatic bioreactors provide environments with lower shear, compared to stirred tank bioreactors [1,10,18,19].
However, there are exceptions to this general behavior. Studies have evaluated cellulase and xylanase production using different strains of Aspergillus sp., comparing the performance of pneumatic and stirred tank bioreactors [[20], [21], [22]]. Typically, higher enzyme yields were achieved using the pneumatic bioreactors (airlift and bubble column), which could be attributed to the low shear environments in pneumatic bioreactors. However, enzyme production by filamentous fungi is favored by a dispersed hyphal morphology. Consequently, higher enzyme production may be induced by the higher shear rate in pneumatic bioreactors, because exposure of the fungi to the shear environment leads to a predominantly dispersed morphology. On the other hand, when Aspergillus sp. grows with pelleted morphology, production of citric acid may increase [23].
The exceptions mentioned above regarding the effects of shear levels in pneumatic and stirred tank bioreactors have also been observed in other studies. Siedenberg et al. [24] cultivated Aspergillus awamori in stirred tank and airlift bioreactors, using wheat bran as carbon source and inductor of xylanase production. Analysis of fungal morphology showed pelleted growth in the conventional stirred tank bioreactor, while filamentous mycelium formation was observed in the airlift bioreactor, which could have been due to a higher shear environment in the latter device.
Cerri and Badino [17] correlated the production of clavulanic acid by Streptomyces clavuligerus with the average shear rate in 4 L stirred tank and 6 L concentric-duct airlift bioreactors. For the same initial oxygen transfer condition, the maximum value of the fermentation broth consistency index (Kmax), which is a rheological parameter related to the morphological structure of the mycelium, was lower for cultivation in the concentric-duct airlift bioreactor, compared to the stirred tank bioreactor. It was found that a higher shear rate was associated with greater fragmentation of the bacterial hyphae and a lower consistency index of the broth, indicating higher shear rates in the pneumatic bioreactor.
Jesus et al. [25] evaluated the hydrodynamics and oxygen transfer in stirred airlift and stirred tank bioreactors operated with xanthan gum solution, at specific air flow rates (φair) ranging from 0.5–1.5 vvm. At the same agitation speed (N), the values of the volumetric oxygen transfer coefficient (kLa) and average shear rate () were higher for the stirred airlift bioreactor, due to the additional liquid circulation flow pattern observed in this bioreactor. However, the shear level depended as much on the presence or absence of mechanical agitation as on the operating conditions (N and φair).
These previous findings question the established protocol for evaluation of the operation and performance of bioreactors, since they suggested that may not be an ideal parameter to use for characterization of shear conditions in bioreactors. In fact, adoption of a maximum shear rate () would seem to be more suitable for the purpose of comparison among different bioreactor models, since it describes the worst condition to which a microorganism could be exposed. Depending on the value, a single exposure to this condition could be sufficient to cause irreversible damage to the cells. The represents a local value associated with a particular location inside the bioreactor. In stirred tank bioreactors, the region around the impeller exhibits the highest shear rate [26], while in airlift bioreactors, this worst shear condition is found in the bottom region, around the sparger holes [[27], [28], [29]]. Nonetheless, despite the effect that has on cells, several other factors may affect the resistance of cells to shear, such as cell size and the mechanical resistance of the cell wall.
The shear conditions in different pneumatic bioreactors were evaluated by Thomasi et al. [6], who performed Streptomyces clavuligerus cultivations for clavulanic acid production in 5 L bubble column (BC), concentric-duct airlift (CDA), and split airlift (SA) bioreactors. Using the relation between the maximum value of the fermentation broth consistency index (Kmax) and the average shear rate (), proposed by Cerri and Badino [17], Thomasi et al. [6]. obtained the following order for the average shear rate: . The value of appeared to be related to the capacity of the bioreactor to produce an internal circulation, since the lowest values were found for the bubble column bioreactor, which had no well-defined liquid circulation pattern.
The evaluation of can be performed using computational fluid dynamics (CFD) to solve conservative equations (continuity and momentum), with calculation of the flow fields and velocity profiles enabling estimation of local and average shear rates for different fluids and operating conditions. CFD has been extensively applied for evaluation of the performance of pneumatic bioreactors, mainly by analyzing variables such as liquid velocity [[27], [28], [29], [30], [31], [32], [33], [34], [35], [36], [37], [38], [39], [40], [41], [42]], global and local gas hold-up [[27], [28], [29], [30],32,33,[35], [36], [37], [38], [39], [40], [41],[43], [44], [45]], and volumetric oxygen transfer coefficient [28,31,38,43]. However, few studies using CFD to evaluate the shear conditions in pneumatic bioreactors are reported in the literature [[27], [28], [29],31,37,44]. Recently, Esperança et al. [46] showed that CFD was a suitable tool for estimation of the average shear rate () in pneumatic bioreactors, obtaining values for this parameter within an expected range. In the present work, the aim was to evaluate and compare the average shear rate () and maximum shear rate () in airlift and bubble column bioreactors, as well as to obtain a relation between and the sparger characteristics of pneumatic bioreactors.
Section snippets
Bioreactors and experimental setup
In this study, CFD was employed to evaluate the shear rate in the following types of pneumatic bioreactor: bubble column (BC), split airlift (SA), and concentric-duct airlift (CDA).
The pneumatic bioreactors evaluated had two different working volumes: 5 and 10 L. Four different geometries of 10 L square cross section airlift bioreactor were simulated, comprising two concentric-duct airlift (CDA) models and two split airlift (SA) models, based on the work of Esperança et al. [47]. Three
Average shear rates in the pneumatic bioreactors
The average shear rate values obtained from the CFD simulations were calculated by adopting a volume averaging procedure for the spatial distribution of the shear rate throughout the bioreactor volume. Fig. 4 presents the values for the 5 L pneumatic bioreactors. For the bubble column bioreactor, ranged from 11–17.1 s−1, while for the split airlift and concentric-duct airlift bioreactors, this parameter varied from 12.7–27.3 s−1 and from 15.6–25.3 s−1, respectively. The loop (airlift)
Conclusions
Computational fluid dynamics (CFD) was used to evaluate the average shear rate () in three different pneumatic bioreactors (bubble column, concentric-duct airlift, and split airlift) operated with different fluids. At the same specific air flow rate, the airlift bioreactors (concentric-duct and split) exhibited higher than the bubble column bioreactor, due to the liquid circulation pattern observed in the former devices.
The average shear rate () in the pneumatic bioreactors
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
CRediT authorship contribution statement
Mateus N. Esperança: Conceptualization, Methodology, Software, Visualization, Writing - original draft, Writing - review & editing. Caroline E. Mendes: Investigation. Guilherme Y. Rodriguez: Investigation, Software. Marcel O. Cerri: Conceptualization, Writing - review & editing. Rodrigo Béttega: Methodology, Writing - review & editing. Alberto C. Badino: Conceptualization, Supervision, Writing - review & editing, Funding acquisition.
Acknowledgments
The authors are grateful for the financial support provided by the Human Resources Program of the Brazilian National Agency of Petroleum, Natural Gas, and Biofuels (PRH/ANP-44), the National Council for Scientific and Technological Development (CNPq, grant 478472/2011-0), the São Paulo State Research Foundation (FAPESP, grants 2011/23807-1 and 2012/17756-8), and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES, Finance Code 001).
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