Studies of the shear protective effects of Pluronic F-68 on wild carrot cell cultures

Abstract

Numerous studies have confirmed that Pluronic F-68 can effectively protect mammalian cell cultures grown in intensely agitated bioreactors from the damaging effects of hydrodynamic shear. Like mammalian cells, plant cells exposed to intense shear conditions are also severely damaged. However, the possibility of shear-protection for plant cells by Pluronic F-68 (PF-68) is seldom addressed. This work addressed this deficiency and investigated the shear-protective properties of PF-68 on wild carrot cultures ( Daucus carota), using a two-level, three-factor (2 3) factorial design. The aim of the experiments was to determine whether PF-68 could provide similar protection for plant cells compared to animal cells, using a novel shearing apparatus [Chem. Eng. Res. Des. 79 (2001) A8]. The three factors involved in the study were impeller speed (300, 400 rpm), exposure time (3, 4 h) and the concentration of the proposed shear-protective agent—PF-68 (0.05, 0.1%, w/v). As biological responses of plant cells change exponentially with shear stress and time, narrow ranges of the speed and exposure time were necessary to ensure the linearity of the results. The biological responses of the plant cells were quantified as relative mitochondrial activity, aggregate size and membrane integrity. Student’s t-tests confirmed a significant improvement in the biological responses for the cell cultures grown in a medium supplemented with PF-68. Cell breakage was reduced considerably for cells grown in medium supplemented with PF-68. The principal variables affecting growth were agitator speed and time. In addition, the mitochondrial response showed strong interactions between concentration, shear and exposure time. These interactions implied that the concentration of PF-68 providing optimal protection may depend on the shear conditions.