Note on spectrophotometric determination of proteins in dilute solutions

Abstract

In order to describe a relatively novel mechanism associated to heavy metal resistance in Euglena gracilis, the osmotic responses during cadmium accumulation were analyzed. Several physiological/biochemical parameters were assessed in E. gracilis exposed to 50 and 200 μM Cd2+, concentrations well within the range found in polluted soils and aquatic environments. The Cd2+ accumulation attained after only 24 h exposure induced marked increases in both the intracellular water volume and cellular size, which were maintained for several days; these changes correlated with (i) an increased intracellular osmolarity (i.e. osmotic pressure) driven by a generalized increase in the osmo-metabolites (trehalose, phosphate-molecules, amino acids, thiol-molecules, betaines and polyamines) content; and (ii) the triggering of antioxidant defenses (increased GSH/GSSG ratios and GPx, GR and APx activities). In contrast, no changes in intracellular volume, cellular size and antioxidant status were observed under hyperaccumulation of Zn2+. Furthermore, E. gracilis cultured in a hypoosmotic medium promoted greater Cd2+ accumulation and water volume, compared to a hyperosmotic medium. These changes were significantly decreased by the aquaporin inhibitors Hg2+ and pentamidine. The results suggested that the cell responses to Cd2+ and osmotic stress share biochemical mechanisms in this algae-like protist, in which Cd2+ accumulation closely correlates with the intracellular water volume changes in an oxidative stress-mediated process.