Effect of copper on pro- and antioxidative reactions in radish (Raphanus sativus L.) in vitro and in vivo

The generation of superoxide radicals, lipid peroxidation (as measured by malone dialdehyde formation) and the activity of selected antioxidant enzymes (superoxide dismutase, catalase, ascorbate peroxidase) were assessed in radish (Raphanus sativus L.), in response to elevated concentrations of copper ions in the culture medium in vitro and in vivo. Experiments were performed on 7-day-old seedlings and 5-week-old calluses grown on media supplemented with CuSO4 in concentrations of 10, 100 and 1000 μМ. The exposure to elevated Cu concentrations in the medium significantly reduced both callogenesis and the proliferation of radish calluses in vitro. Cu treatment resulted in the increased generation of the superoxide radical (O2−) in radish seedlings and calluses indicating the occurrence of oxidative stress in radish cells, whereas the level of lipid peroxidation (LPO) remained unchanged. Both in calluses and in radish seedlings in vivo, the relative level of oxidative stress was maximal at micromolar Cu concentrations and became attenuated with increasing Cu concentrations. Stronger oxidative stress occurred in the radish seedlings in vivo, compared with radish calluses in vitro. The observed lower sensitivity of calluses to Cu-induced oxidative stress and their ability to proliferate upon exposure to Cu concentrations of up to 1000 μМ demonstrate the potential of in vitro cell-selection to obtain metal-tolerant radish plant lines.