Determination of shear stress thresholds in toxic dinoflagellates cultured in shaken flasks: Implications in bioprocess engineering

Marine dinoflagellates are potentially important innovative sources of high-value toxins in biomedical, toxicological and chemical research programs. However, little is known about the difficulties related to dinoflagellate cultures. In this article, we demonstrate that the shear sensitivity of cells may be one of the main causes. The red-tide Protoceratium reticulatum, a producer of yessotoxins, was used to examine the effect of hydromechanical shear stress associated with intermittent fluid agitation on cell growth. Shaken flasks, widely used in biotechnological process research, were used as model bioreactors, as hydrodynamic shear stress is relatively easy to quantify in them. Intermittent turbulence regime was characterized by three key operating variables: shear stress, cycle time or shaking frequency, and fraction of time shaken per agitation cycle. The light/dark cycle was also used as another variable. Cell damage depended on the combination of the above-mentioned variables. A damage threshold was observed at an average shear stress of approximately 0.16 mN m−2 (equivalent shear rate of 0.12 s−1). Cell damage from exposure to average detrimental shear stress was also shown to be greater in the dark than in the light period. Preliminary experiments demonstrated that dinoflagellates are also much more sensitive to bubbling than the majority of common fragile microalgae. Although slight toxicity of Pluronic F-68 was observed at a concentration of 0.05% (w/v), this protective medium additive considerably reduced cell breakage. On the other hand, no cell adaptation to stronger shear stress was observed. Finally, the implications of the proposed approach for the hypothetical mass culture of dinoflagellates in bioreactors were also thoroughly assessed.