Alleviation of copper rhizotoxicity by calcium and magnesium at defined free metal-ion activities

The common "hardness cations", Ca and Mg, are at least partially capable of alleviating the toxicities of metal ions, but quantitative data for their effect on Cu rhizotoxicity are lacking. We used 2-d root elongation of wheat (Triticum aestivum L,, cv. Yecora Rojo) in a simple medium (greater than or equal to 0.2 mM CaCl2) as a sensitive bioassay for examining how Ca, Mg, and pH affect Cu rhizotoxicity. A fourfold increase in solution Car from 1 to 4 mM showed a slight alleviation of toxicity, but the effect could be completely ascribed to changes in ionic strength and thus in Cu2+ activity. Subsequently, a 25-fold range in Ca, (17-fold range in Ca2+ activity) was found to improve Cu-inhibited root elongation by some 30 to 35%. Increases in solution Mg2+ activity were equally or slightly more effective than comparable increases in free-ion activity [denoted by parentheses, i,e., (Ca2+)] of Ca in alleviating Cu stress, an unexpected result because of reports of selective binding of Ca by apoplastic sites. Neither cation alleviated Cu rhizotoxicity as profoundly as they do with other metals such as Al. Lowering pH from 6.5 to 5.5 alleviated Cu stress slightly; a further decrease to pH 4.5 was more effective. Finally, seedlings were reared en masse in nine treatments selected from the previous root elongation studies, and root tips were analyzed for operationally defined apoplastic and symplastic Cu, Growth inhibition was only weakly correlated with symplastic Cu (or with apoplastic or total Cu), and the results suggested that Ca and Mg (but not H) afford some physiological protection against Cu that is not explained by simple competitive inhibition of Cu accumulation in the apical tissues.