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- Mixing efficiency on plant cell growth and proteinase production in a stirred tank reactorPublication . Raposo, Sara; Lima-Costa, Maria EmíliaMixing efficiency is one of the most significant factors in bioprocess productivity. The major role of agitation is to improve broth homogenization, mass and heat transfer inside the bioreactors.
- Focussed beam reflectance measurement (FBRM) monitoring of particle size and morphology in suspension cultures of Morinda citrifolia and Centaurea calcitrapaPublication . Jeffers, Paul; Raposo, Sara; Lima-Costa, Maria Emília; Connolly, Patricia; Glennon, Brian; Kieran, PatriciaLaser light scattering technology, as applied in the Lasentec focussed beam reflectance measurement (FBRM) system, was used to characterise two morphologically dissimilar plant cell suspension cultures, Morinda citrifolia and Centaurea calcitrapa. Shake-flask suspensions were analysed in terms of biomass concentration and aggregate size/shape over the course of typical batch growth cycles. For the heavily aggregated C. calcitrapa, biomass levels [from 10-160 g fresh weight (fw) l-1)] were linearly correlated with FBRM counts. For M. citrifolia, which grows in unbranched chains of 2-10 elongated cells, linear correlation of biomass concentration with FBRM counts was applicable in the range 0-100 g fw l-1; at higher levels (100-300 g fw l-1), biomass was non-linearly correlated with FBRM counts and length-weighted average FBRM chord length. For both cell systems, particle morphology (size/shape) was quantified using semi-automated digital image analysis. The average aggregate equivalent diameter (C. calcitrapa) and average chain length (M. citrifolia), determined using image analysis, closely tracked the FBRM average chord length. The data clearly demonstrate the potential for applying the FBRM technique for rapid characterisation of plant cell suspension cultures.
- Effects of the hydrodynamic environment and oxygen mass transfer on plant cell growth and milk-clotting protease production in a stirred-tank reactorPublication . Raposo, Sara; Lima-Costa, Maria EmíliaIndustrial-scale plant cell based bioprocesses are limited because of technological challenges associated with mass transfer and mixing in the heterogeneous broth, which typically displays non-Newtonian characteristics. Cultures of Centaurea calcitrapa cells, presenting milk-clotting activity, were established in a 7-L stirred-tank reactor (STR), keeping the initial mass transfer conditions constant, to gain a better understanding of the effects of the hydrodynamic environment and mass transfer on protease production. Cell suspensions grown in an STR equipped with one single marine propeller or two Rushton turbines showed the highest protease activities and biomass production, corresponding to the adequate mass transfer achieved under the tested bioreactor conditions. The key factors improving milk-clotting protease production in a bioreactor were an efficient mass transfer and good bulk mixing without the formation of stagnant zones, while a compromise had to be established in relation to the hydrodynamic shear conditions.
- Use of focussed beam reflectance measurement (FBRM) for monitoring changes in biomass concentration (Erratum)Publication . Whelan, Jessica; Murphy, Eilis; Pearson, Alan; Jeffers, Paul; Kieran, Patricia; McDonnell, Susan; Raposo, Sara; Lima-Costa, Maria Emília; Glennon, BrianThe potential of focussed beam reflectance measurement (FBRM) as a tool to monitor changes in biomass concentration was investigated in a number of biological systems. The measurement technique was applied to two morphologically dissimilar plant cell suspension cultures, Morinda citrifolia and Centaurea calcitrapa, to a filamentous bacteria, Streptomyces natalensis, to high density cultures of Escherichia coli and to a murine Sp2/0 hybridoma suspension cell line, 3–2.19. In all cases, the biomass concentration proved to be correlated with total FBRM counts. The nature of the correlation varied between systems and was influenced by the concentration, nature, size and morphology of the particle under investigation.
- Rheology and shear stress of Centaurea calcitrapa cell suspension cultures grown in bioreactorPublication . Raposo, Sara; Lima-Costa, Maria EmíliaCentaurea calcitrapa suspension cultures were grown either in Erlenmeyer flasks or in a mechanically stirred bioreactor. Its rheological behaviour, when fitted to the Oswald–de Waele model (power law), showed pseudoplastic characteristics in both cases. The flow behaviour index (n) decreased over the course of a growth cycle and the consistency index (K) increased, reached a value of 1.81 N sn m−2 run on 2 l bioreactor. Bioreactor cultivation of C. calcitrapa cells at different agitation rates (30, 60, 100 and 250 rpm), highlighted the influence of shear forces on cell viability loss (90–34%) and phenol accumulation (74–140 μg l−1), due to increased stirring speeds. Analysis of these results suggests that this cell line is shear-sensitive. An empirical exponential correlation was defined between apparent viscosity and biomass concentration, under the studied conditions, giving the possibility to estimate the prevailing broth regime and to optimize bioreactor design.